Week 20 of SIP: Last Posting

Name: Hardina Bte Hamzah
Topic: Cytogenetics

Structural Abnormalities of Chromosomes

When chromosomes have replicated and are waiting to be separated into 2 daughter cells, chromosome breakage is frequently accompanied by exchange material from one chromatid to another. The exchange of material occurs during the pachytene period of the first meiotic division and is a mechanism built into the reproductive cycle to ensure the mixing of the gene pool. The exchange is called crossing over and it occurs when paired homologs form chiasmata. The abnormailties occur when the chromosomes break at non-homologous sites leading to unequal exchanges. 2 major classes of detectable rearrangements are rearrangements within a single chromosome and rearrangements involving more than 1 chromosome.

Examples of rearrangements within a single chromosome:

Deletion is a loss of chromosomal material. An interstitial deletion can arise from unequal crossing over at meiosis. Breaks can occur in 2 homologs at slightly different positions on the chromosomes and the pieces can be reconnected to the alternate chromosome. One chromosome could lose material while the other could gain material. This kind of mechanism is more likely to occur. Cri-du-chat syndrome is one of the examples of syndromes resulting from deletion.

Duplication occurs when a portion of one homolog may be present in duplicate. A duplicated segment may be inserted in the same order as the original segment. However, we cannot identify which piece was original. It can also become reversed. Duplication in the same order which also can be known as tandem duplication may probably results from unequal crossing over in meiosis or from a rearrangement between 2 chromatids during mitosis. Any duplication represents a trisomic state for whatever gene loci are present on the affected segment.

Inversion is a two-break structural irregularity that affects a single chromosome. The segment between the 2 breaks rotates 180 degrees and reattaches to the points of breakage. There are 2 types of inversion: Paracentric and Pericentric. In paracentric inversions, the centromere is not included in inverted segment and the arm ratio of the chromosome remains the same. In pericentric inversions, the inverted portion does include the centromere and the arm ratio may be changed.

Examples of rearrangements involving more than 1 chromosome:

Translocation is an exchange of genetic material between 2 or more non-homologous chromosomes. It is an abnormal mechanism but it can occur when 2 or more chromosomes break at the same time. Broken chromosome ends are regarded as sticky and can attract the cellular enzymatic repair service. Usually broken ends stay close to each other and are reunited. However, it is possible to mismatch the broken ends. The carrier of a balanced translocation may pass on the only part of the translocation which can result in genetically unbalanced offspring.

Dicentric chromosome possesses 2 centromeres and it is a result of the joining of homologous chromatids to form an isochromosome. An isochromosome is a chromosome in which the arms n either side of the centromere are morphologically identical ad bear the same gene loci. Dicentrics may cause problems during cell division, however, if the 2 centromeres are far apart on the derivative chromosome. If both centromeres are active, they can be drawn to opposite poles of the spindle resulting in formation of an anaphase bridge. Anaphase bridge is a chromosome that makes a bridge between 2 daughter cells at anaphase. When this happens, the dicentric can be left outside both daughter nuclei as they form or can break apart and cause loss or gain of chromosomal material.

Insertion is an addition of genetic material between 2 normally adjacent sections of a chromosome. The additional material could be a duplication of a section of the same chromosome but is more likely to come from a different chromosome. Insertions are difficult to identify unless there is a complementary deletion in the cell.

WEEK 19 Posting

Architect system analyser (ci8200)

Picture taken from: http://www.drdangslab.com/images/eqiupments/ci8200.jpg



Introduction

The modular design of the ARCHITECT family of analysers allow multiple processing modules which perform all sample processing activities, to be physically joined to form a single workstation or system. These ARCHITECT systems can be configured to process sample using potentiometric and photometric methoeds and/or chemiluminescent microparticle immunoassay (CMIA) methods.

The ARCHITECT ci8200 analyser is composed of two modules, which is c8000 and i2000. C8000 module is using the potentiometric and photometric methos while the i2000 module uses CMIA method.

Here are the majority tests carried out the ARCHITECT analyser and their reference range:


1. Free Thyroxine (FT4) 10.0-20.0pmol/L
2. Total Triidothyroninn (Total T3) 1.2-2.3 nmol/L
3. Anti-TPO <5.6IU/ml
4. Anti-Tg <4.11u/ml
5. Alpha-fetoprotein (AFP) 1.0-10.0ug/L
6. Carcinoembroynic Antigen (CEA) 0.0-5.0ug/L
7. Vitamin B12 (B12) 132-835 pmol/L
8. Folate (FA) 7.40nmol/L
9. Ferritin 30-323ug/L M (0-30Y) 32-294ug/L M (31Y & Above) 7.6-179ug/L F (premenopausal) 182-339ug/L F (postmenopausal)
10. Syphilis non-reactive
11. HBsAg non reactive
12. HbeAg non-reactive
13. Anti-HCV non-reactive
14. Anti-HAV IgM non-reactive
15. HAV AB non-reactive
16. Anti-HBs non-reactive
17. HIV Ag/Ab Combo non-reactive
18. Kappa light chains 1.40-3.8g/L
19. Lambda light chains 0.90-2.50g/L
20. Immunoglobulin A 0.8-4.0g/L
21. Immunoglobulin G 5.0-15.0g/L
22. Immunoglobulin M 0.3-3.0g/L
23. Rheumatoid Factor <20.0 IU/L
24. Transferrin 1.74-3.64g/L Spectrophotometric method
25. Iron F 11.27 umol M 11.29umol/L

The analyser uses these three methods to measure the concentration of analytes in sample.

1. Photometry method

Phtometry method uses
-End point assay are reactions that reach an equilibrium and at that time there is little or no further change to the absorbance readings. During this equilibrium time, the system measures the absorbance readings used for calibration and calculating results.

-Rate assay are reactions that measure the constant change in absorbance over time. The system performs readings several times during this reaction, calculates absorbance change (activity), and then uses the readings to calculate results.

2. Potentiometry (measures electrical potential in sample)

-The c8000 modules uses an ICT (integrated chip technology) modules to measure potentiometric assay (eleytrolytes). ICT methodology uses the solid state ion-selective electrodes contained in a single chip that reduces the maintenance require to perform electrolytes measurements.

3. Chemiluminescent technology

-During sample processing, microparticles (paramagnetic microparticles coated with capture molecules) are mixed with the sample binds to the corresponding capture molecules on the microparticles, forming the immune complex.
-After incubation, a magnet attracts the paramagnetic microparticles (bound to the surface analyte) to a wall of the reaction vessel. The reaction mixture is washed to remove unbound materials, further processing can now take place.
-Detection of ther eaction mixture in the reaction vessel is accomplished with a chemiluminescent acridinium-labeled conjugate. The conjugate binds to the immune complex to complete the reaction mixture.
-After incubation, the reaction micture is washed to remove unbound material. The pre-trigger solution is added and a background read is taken.
-Pre-trigger performs the following functions: - Create an acidic anvironment to prevent early release of the energy(light emission). Helps to keep microparticles from clumping. - Splits acridinium dye off the conjugate bound to the microparticle complex. This prepares the acridinium dye for the nest step.
- The system then adds Trigger solution to the reaction mixture. The acridinium alkaline solution. This causes the chemiluminescent reaction to occur. N-methylacridne is formed and releases energy (light emission) as it returns to its ground state. The CMIA optical assembly measures the chemiluminescent emissin (activated red) over a predefined time perioed to quantitate the analyte concentration or to determine qualitative interpretations for index cut-off assay.

Specimen

-Human serum (including serum collected in serum separator tubes) or plasma collected in sodium heparin, lithium heparin separator tubes, or potassium EDTA anticoagulant tubes may be used. Other anticoagulants have nor been validated for use with ARCHITECT.

Method
1. Specimen are spun via the centrifuge for 5minutes.
2. The cap of the specimen tube is removed.
3. Specimen placed on the ARCHITECT rack.
4. Rack loaded into the ARCHITECT.
5. Test are run.

So that's all for my last posting.

Thanks for reading.

Dyana
0605169B

Week 18-my final entry =)

Subject title: Clinical Chem/Micro
Name of test: Urea Breath Test

Principle:
-This test is carried out for diagnosis of Helicobacter pylori (H pylori) infection
-H pylori is a gram-negative bacillus which is highly motile due to its multiple unipolar flagella

Retrieved 21 October, 2008, from http://images.google.com.sg/imgres?imgurl=http://www.science.org.au/nobel/2005/images/invasion.jpg&imgrefurl=http://www.science.org.au/nobel/2005/05diagram4.htm&h=329&w=321&sz=30&hl=en&start=2&um=1&usg=__Xjdem48HW8H_C711OrWmzPSMn7g=&tbnid=0Tjg0gylcn5OMM:&tbnh=119&tbnw=116&prev=/images%3Fq%3DH%2Bpylori%26um%3D1%26hl%3Den%26sa%3DN

*thread-like things=flagella
-They produce the enzyme urease which converts urea to ammonium and bicarbonate
-Some of the diseases caused by H pylori infection are:
1. Gastritis and dyspepsia
2. Stomach ulcers
3. Duodenal ulcers
4. Stomach cancer
5. Lymphoma

-The UBIT Tablet taken by the patient contains 100 mg of 13 C-Urea
-If the patient has Helicobacter pylori infection, H. pylori will produce the enzyme urease. The enzyme urease will convert the 13 C-Urea to 13 CO2 and NH4+
-The 13 CO2 is then absorbed in the blood, diffused into the blood and exhaled
-In our laboratory, we use the 13 CO2 Urea Breath Analyzer UBiT-IR300 by Otsuka electronics to analyse our UBT samples
-The machine will measure the 13 CO2 in both the *pre-dose and post dose breath samples
-The difference of the 13 CO2 levels in both breath samples will also be calculated
-This measurement is done via infrared spectroscopy

Materials:

2 UBT bags
1 UBIT tablet
100mL of water


Method:
Preparation for the test:
Patient must:
1.Fast for 8-9 hours before the test
2.Stop taking Proton Pump Inhibitors one week before the test
3.Stop taking antibiotics 4 weeks before the test

During the test:
1. The patient has to breathe into the “Pre-dose” bag without taking the UBIT tablet.
2. Within 5 sec, the patient has to swallow the UBIT tablet on an empty stomach with 100 mL of water. The tablet is not to be chewed, crushed or dissolved.
3. After taking the tablet, the patient has to lie down on his left side for 5 minutes.
4. Then, he has to remain seated for another 15 minutes.
5. After that, the patient has to breathe into the “Post-dose” bag.
6. These two bags with the request form will be sent our lab for analysis.


In the lab:
The medical technologists will:
1.Check the labels
2.Insert “Pre-dose” bag into the “Pre-dose” inlet
3.Insert “Post-dose” bag into the “Post-dose” inlet
4.Verify results

Results:
A value of 2.5% or higher indicates that the patient is positive for H.pylori infection

*pre-dose: sample before taking tablet

*post-dose:sample after taking tablet

Nur Farhana
0604834B

week 17

Well for the past week, i was assigned to do shift work.
i had to work from 10am till 7pm. the slot was either for Haematology section or CP.
CP stands for Corporates and Poly section.
For haematology, the workflow remain the same. i will just have to assist the medtech that is posted there and cover the section from 5pm-7pm or until the next shift staffs are in.

However i was new to CP section. So i was asked to observe the workflow first.
The specimens from the polys will arrived at the lab around 10.30am. That will be the first batch of specimens. In total, 3 batches of specimens are send to the lab within the expected timings. Upon receiving the specimens, the poly clerk will have to check the specimen sent, wether it matches will the ones indicated in the forms. Once checked, the clerk will have to 'ech' the specimen into our system. If 'ech' is not done, the specimen cannot be run as the the machines will not be able to recognise them. After which the poly medtech will take over. He/ She will then have to process the specimen according to the test requested.
Usually the bloods are sent in plain tubes and fluoride tubes.
For plain tubes, we can load it into MPA ( a machine in chemistry) directly. For fluoride tubes however, we have to spin them using centrifuge before loading them into either SWA or COBAS for processing.
The tubes are then archive into a special rack for easy indentification.
After the processing of the first batch is done, we have to check for any test that are not run. We do so by checking the incomplete log. In this system, we can check for any outstanding test that are not yet done.
If there is indeed any test that are not run, we have to find the specimen and run it.
That's for poly specimens.

For corporate specimens, the workflow remains the same. The only difference is that the specimen are not 'ech' into our system but are ordered as corporate screening specimen. What we do here is, we ordered the tests that are required like how we do order entry but the location for the test are change into their respective companies.
The workflow henceforth are similar to the polys.

After all the specimens are received and processed, we have to check the incomplte log again in case we missed out any test. If everything is smooth sailing then we can print the corporates results which are then reported back to the companies.

So that's basically the roles of a CP medtech.
till then...

sutiana
0604651j
to1

Week 16

hello! Since the past two weeks, i have been assigned to be at the special stains station, alongside with a senior medical technologist. The special stains station consist of a machine responsible for stains such as GMS(Gomori's Methenanine Silver Nitrate), PAS(Periodic Acid Schiff), PASD(Periodic Acid Schiff Diastase), Fe(iron), Alcian Blue, Giemsa, Steiner, Retic.

Picture taken from: http://www.ventanamed.com/

The Nexes Special Stain machine is barcode-driven and has optimizes protocols to help ameliorate productivity and consistency. The tray in the machine can hold up to 20 slides while the reagent tray can take up to 25 reagents.

1) The sections are fished from the floatation bath and onto the slide.
2) The biopsy number and the name of the special stain required is labeled on the slide.
(ie. 08/12345 GMS)
3) The slide is heated on the hot plate.
4) The barcode label is keyed in and the protocol of the test required for the slide is chosen.
5) The label is pasted on the slide.
6) The slide is sent for dewaxing.
7) The slide is placed in the slide tray in Nexes.
8) The section on the slide is hydrated with a wash solution to prevent drying up of the section and to prevent unwanted solution in the machine to dirty parts of the slide.
9) The reagents required are loaded.
10) The liquid coverslip solution is filled up if it is insufficient.
(The liquid coverslip acts as a temporary protection while the slide is still in the machine after staining)
11) The RUN button is clicked
12) The run time will be shown
13) When staining is done, the reagents are removed from the tray.
14) The slide is removed from the slide tray and sent for dehydration(70%, 80%, 95%, absolute alcohol)
15) The slide is cleared, by dipping it into xylene.
16) The slide is mounted.


So far, i have performed two stains manually, without the help of the Nexes Special Stain machine. The stains are PAS and PASD. PAS stain is used to stain carbohydrates. The difference between the two stains is, PASD uses diastase to digest glycogen. This stain is used to differentiate glycogen from other carbohydrates. Under microscopic examination, the tissues stained with PAS will consist of clusters of glycogen, stained dark pink / magenta.

1) The section of interest is fished onto the slide.
2) The slide is heated on the hotplate for 3 minutes.
3) Dewax the slide and bring the sections to water.
4) For PASD stains, treat the sections with thawed diastase for 30 minutes and subsequently, wash the slide with water.
5) For PAS stains, treat it immediately with 1% aqueous periodic acid for 5 mins.
6) Treat the slides with Schiff's reagent for 3 mins, until sections turn light pink.
6) Wash the slides in water
7) Counterstain the nuclei with alum haematoxylin for 1 min.
8) Blue the slides in water
9) Dehydrate the slides by dipping them into 70%, 80%, 95&, absolute alcohol.
10) Clear the slides by dipping them into xylene
11) Mount the slides

PAS stain- note the clusters of magenta

Picture taken from: http://www.path.utah.edu/casepath/PM%20Cases/PMCase6/ALL6_files/image046.jpg

Nurathirah
0606561I

Week 15 Of SIP

Topic: Processing Section

To all my muslim friends, Selamat Hari Raya! And to all my non-muslims friends, hang in there...5 weeks left. Sorry fot the late blog entry. My computer is not working well these days.

For the last 5 weeks, I have been attached at the processing section. Some feels that the processing section is rather simple yet boring job. But I feel that it is the MOST important section in the laboratory. Every specimen will have to undergo the processing section before the requested tests are performed. Here’s what I have learnt I have pick up along the way…

When a test is requested by physicians, specimens are collected in appropriate tubes/ containers and sent to the lab together with a request form. In the lab that I am attached to, there are four types of tests forms. Such includes the

1. External form-specimens will be sent to external laboratory upon request as test are not available in our lab.
2. Interlaboratory form- request for tests done in the clinical lab.
3. Microbiology lab form-request for test done in the micro lab.
4. Histopathology form-request for test done in histo lab.

Thus upon arrival of specimens to the processing section, it will be sort as according to the forms. Those interlaboratory tests are then further arranged. The interlaboratory test specimens usually come from wards, specialist clinics, accident and emergency section. Those ‘URGENT’ cases from any of the above department will be prioritized. All A&E tests and MICU and SICU cases are also prioritized, followed by clinics, and wards.

The laboratory admin clerk will then order tests requested in the LIS and print out the barcode for the tests requested. The barcode labels are pasted in the sample tubes. When pasting the barcode labels, one must check that the label which was already pasted by the physician tally with the name on the requested form and that the barcode labels are pasted on the right sample tubes for the right tests. E.g. Test for full blood count, specimen must be placed in the EDTA tube and not plain tube.

Medical Technologies must ensure that barcodes are labeled neatly and appropriately so that the analyzers are able to scan the barcode. The specimens are then spin (if required like plain blood tubes) and/or delivered to the respective section.

The processing section is also in charge of rejecting specimens and cancellation of tests. Here are some common reasons that specimens are rejected:
1. blood in EDTA tubes are clotted
2. insufficient specimen
3. specimen leaked
4. empty container sent
5. specimen sent in wrong tube
6. no name label on specimen
7. name/ NRIC mismatch between request form and specimen

Alrite people.Thanks for reading.See you guys soon...

Dyana
0605169B

Week 14:3rd Campus Discussion Week

Name: Nurdyana Binte Abdul Rahman


Test Name: Stool for ova, amoeba & cyst test.


Hello everyone. Hope you guys are coping well. I have observed this test done at the Routine Section. The ova and parasites test is performed to identify intestinal parasites and their eggs, which are called ova, or cysts in patients with symptoms of gastrointestinal infection. Patients may have no symptoms or may not even experience diarrhea, blod in the sool, and other gastrointestinal distress. Identification of a particular parasite indicates the cause of the patient's disease and determines the medication needed to treat it.

Saline wet mount would be the best for detection of helminth eggs and larvae and it is especially good for motile protozoan cysts, which appear refractile. When cysts have been found in saline mount, they should be observed in Lugol iodine stained preparation which will reveal nuclear structure and te presence of glycogen. The nuclei stain a brownish colour and can be easily stained. For an accurate diagnosis, a concentrated method is advised so as to increase the probability of finding the parasites in faecal samples.


Para-pak spin con is a commercial kit for fecal parasite concentration of eggs, larva and protozoa. The picture below is para-pak spin con.






Picture taken from http://www.mdeur.com/products/972200.htm

Sample:


Feces should be collected directly into a clean container. It must not be contaminated with urine, water, or other materials.



Reagents:


-10% buffered solution
-Saline
-Lugol Iodine Solution

Procedure:
1. Stool(0.5g) was emusified with 10ml of saline in a plastic tube
2. 4 drops of CON-Trate Reagent A which helps to enhance the breakdown of fecal aggregrates and mucus therefore freeing parasites.
3. Plastic tube was capped and contents were mixed thoroughly.
4. The mixture were then filtered using the para-pak filter into the centrifuge tubes provided.
5. The filtered mixture were span at 2000rpm for 10 minutes.
6. The supernatant were removed.
7. And the sediment were resuspended.
8. Wet mount were performed on the sediment.
9. Then slide were examined microscopically for parasite.


The senior medical technologies will be observing the slide. Presence of bacteria and other microorganism in the stool would indicate patient is normal, thus no ova, amoeba or cyst seen. Presence of ova, amoeba or cyst would indicate an abnormal result. In addition is a sign of parasitic infestation.

Week 14: 3rd Campus Discussion Session

Name: Hardina Bte Hamzah

Topic: Cytogenetics

Title: Amniotic Fluid Set-up

Amniotic fluid is a clear, slightly yellowish liquid that surrounds the unborn baby (fetus) during pregnancy. It is contained in the amniotic sac. The fluid protects the fetus from injury and helps to regulate the temperature of the fetus. It allows the baby to move around so that the muscles and bones develop properly. It also helps the digestive and respiratory system to develop as the baby swallows and inhales and exhales it from his lungs. In addition to various enzymes, proteins, hormones, and other substances, the amniotic fluid contains cells shed by the fetus. These cells contain genetic information that can be used to diagnose chromosomal disorders. The fluid is sent to cytogenetics lab so that the cells can grow and be analyzed.

To collect the amniotic fluid, the obstetrician will insert a very fine needle through the woman's abdomen into the uterus and amniotic sac and withdraws approximately 20ml of amniotic fluid for testing. A local anesthesia will be given to the patient. The obstetrician uses ultrasound images to guide needle placement and collect the sample, thereby minimizing the risk of fetal injury and the need for repeated needle insertions.

Before starting the set-up, the medical technologist has to ensure that the labeling on all petri dishes and tubes is done correctly. The set-up must be done in the Biological Safety Cabinet (BSC) as all specimens must be processed in a sterile environment.

One conical test tube and one slant tissue culture tube are used for each specimen. Before transferring the amniotic fluid from the syringe to the tubes, invert the syringe to ensure that there is adequate mixing of the amniotic fluid. After mixing, equal amount of amniotic fluid is transferred to each tube. The tubes will then undergo centrifugation at 1200 rpm for 10 minutes. The buckets in the centrifuge must be closed before centrifugation starts.

Each specimen will have a record sheet where the medical technologist will record down the volume and appearance of fluid, size and appearance of pellet and the presence of RBCs after centrifugation are done.

A sterile coverslip will be placed in each petri dish using a pair of sterile forceps. The dishes will then be divided into 2 sets. One of the set will have the culture tube. Each set will use a different kind of media. The first set with the culture tube will use complete Chang media and the second set will use the complete Bio-AMF-2 media. Both media consists of antibiotics to prevent any bacteria from growing and contaminating the whole culture. The supernatant will be aspirated out leaving around 0.5ml in the tubes. The Chang media will be added into the culture tube and the Bio-AMF-2 media will be added to the conical tube. The amount of media to be added is depending to the number of dishes used. One dish needs 0.5 ml of media. For example, if the first set consists of 2 dishes and 1 culture tube, 1.5ml is needed to be added into the tube.
After adding the media, the pellet will be gently re-suspended and the cell mixture will be transferred onto the coverslip in the dish. The cell mixture must be confined to the coverslip. A drop of cell mixture will be placed from the conical flask to the last dish of that set. This will act as a backup. 0.5ml of cell mixture will be left in the culture tube. The culture tube will also acts as a backup.

The first set of dishes and culture tube will be placed on one tray and the other set on a separate tray. The two trays will be placed in two separate incubators with independent electrical circuits and gas sources. The cultures will be left inside the incubators and on day 3, the medical technologist will check for cells attachment. Other than that, 1.5ml of media will be added to the dishes and tube. If there is no cell attachment, media will not be added and put on hold for another day. On day 5, the media will be changed and be replaced with a fresh media. After enough cells are grown, the cells will then proceed to tissue culture.

week 14

Subject title: Clinical chem/ Haem
Name of test: G6PD screening test

Principle:

• G6PD test carried out in our lab is only a QUALITATIVE and SCREENING test
  
• G6PD is an enzyme which catalyzes the reduction of NADP+ to NADPH
  
• NADPH keeps glutathione in its reduced form


• Reduced glutathione or GSH protects red blood cells from oxidant damage and hence prevents the lysis of RBCs


• The reagent used in this test contains NADP


• Test principle is as follows:

G6PD (this is supposed to be on top of the arrow in the equation..but i juz can't move it)

Glucose-6-P + NADP+ ----->Gluconate-6-P + NADPH+H+

• The NADP in the reagent will be converted to NADPH if G6PD is present
  
• NADPH produced in the reaction will fluoresce under long-wave UV-light
  
• If there is G6PD deficiency or total absence of it, no fluorescence will be observed
  

Materials:

1. Filter paper

2. Hitachi cup

3. G6PD reagent

4. UV light

5. Incubator

6. Patient's sample in EDTA tube

Method:

1. Pipette 100uL of reagent solution into the hitachi cup which has been labelled with the patient's lab no

2.Add about 2.5uL of patient's sample into the same cup

3. Mix well and pipette 5uL onto the filter paper(1st spot)

4. After 5 mins, pipette another 5uL onto the filter paper(2nd spot) and wait for another 5 mins to pipette again 5uL of patient's sample(3rd spot)

5. Then, put the filter paper into the incubator

6. After 5 mins, observe the filter paper under long-wave UV light

ok the filter paper will look something like this, u would have three spots of blood which are made in 5 mins interval by pipetting 5ul of the patient's sample

Results:

1. Fluorescence indicates presence of G6PD

• usually the 1st spot of blood would not fluoresce well, the 2nd and 3rd ones will

2. Absence of fluorescence indicates the absence of G6PD

Note:

• Deficient cases will be followed up with G6PD quantitative test which is a send-out test


• Negative control is done before our first sample of the day

Nur Farhana Binte Ramlan

0604834B

13 th week

hello guys...

for this entry, i will blog on one of the test that is carried out in microbiogy section.

the test is STOOL CULTURE

Principle Analysis
Acute infectious diarrhoea is caused by a number of different agents: bacteria, viruses and protozoa. Each bacterial agent of infectious diarrhoea has unique pathogenic mechanism that cause a specific symptoms. These symptoms and an adequate patient history are clues that will help the physician catergorise the patient's disease. Antimicorbial therapy can alter the host's normal protective microbiota and thus predispose the patient to bacterail overgrowth that result in diarrhoeal disease caused by organism such as Clostridium difficile, staphylococcus aureus, Candida spp. and Pseudomonas aeruginosa.

For our lab, we routinely cultures stools only for the presence of Salmonella, Shingela and Vibro spp. because of their role as causative agents of bacterial diarrhoea and other enteric diseases.

Organism such as Aeromonas spp., Plesiomonas shigelloides and Yersinia spp. are identified only wen they are present in pure or heavy growth or upon request.

Specimen
stool

• collect the specimen in a sterile, dry, leak-proof container
• select portion containing pus, blood or mucus for culture
• 1 to 2 g of sample is sufficient

Reagents
media

• Alkaline Peptone Broth, AP
• MacConkey Agar, MAC
• Selenite-F Broth, SF
• Thiosulphate-Citrate-Bile salts-Sucrose Agar, TCBS
• TSA with 5% Sheep Blood, BA
• XLD

supplies

• Sterile cotton swabs
• Applicator sticks
• Inoculating loop/straight wire

storage requirements

• store all media (agar plates and broth) at 2 to 8 degree celcius
• leave the media at room temperature overnight before use

Procedures
primary inoculation

• note the appearance of the stools i.e watery, loose, bloody, semi-formed and form
• inoculate stools according to Table 1
• subculture selective broth after 18 to 24 hours of incubation as in Table 2

Culture examination
After incubation, examine all primary and subculture plates for the growth of enteric pathogens.

For Salmonella/Shigella/Vibrio cholerae screenig, examine all the primary and subculture plates for the growth of Salmonella, Shigella and V. cholerae.

For the Vibrio cholerae screening, examine all the primary and subculture plates for the growth of V. cholerae.


Reporting of results


dats all...
i hope you guys learnt something from my posting.
thanks for reading..

sutiana
TG01
0604651J

12th week

Ever since i got back from cytology, i have been at the shaving station in histo. Shaving is the process of removing excess wax and exposing the tissues in the embedded blocks. This will provide a smooth-sailing sectioning of the tissues. Shaving is done using the microtome machine. The difference between sectioning and shaving is, sectioning is done at 4-5 microns while shaving is done at 20 microns.

In addition, the person at the shaving station will be responsible for placing the tissue blocks which require decalcification into RDO. RDO is actually an acid which will decalcify the tissues. I have actually come across tissues which i cannot shave at all because it is too hard. For tissues of that nature, it will have to be placed in RDO even before it is shaved. For other tissues which do not require decalcification, they are placed in a detergent after shaving. This detergent will soften the tissue surface to the perfect level to which that it will be easier to section it. These blocks are placed in the detergent for five minutes and then washed under running water. If the blocks are placed in the detergent longer than five minutes, the staining of the sections on the slides will be compromised. Once, it was left in there for more than 5 minutes, blobs of stains were seen on the slides. This is perhaps due to chemical interactions. The detergent which has penetrated into the tissues will interact with the chemicals from the H&E stains.

There are many things to look out for during shaving. Firstly, score lines. Score lines are lines that appear on the surface of the tissue blocks. These lines are caused by the blunt blades. When score lines are observed, the blades must be removed immediately. Then, the blocks will be shaved again until the lines have disappeared. It is very important to ensure that after shaving a block which requires decalcification, the blade is changed to shave the next block. This is because, obviously, the blade will be blunt after shaving the hard tissue from the block which requires decalcification. If the blunt blade is used to shave a tissue block containing small tissues, it is hard to remove the score lines without taking the risk of losing the tissue after continuous shaving. Thirdly, removal of sutures and staples. Sutures are what seemed to be black threads used during an operation. Amazingly, sutures and staples are used by the surgeons during the operation for the orientation of the organs. Orientation of the organs aid in the trimming of the organs by the pathologists. Sometimes, i believe, unknowingly, when the pathologists trim the tissues into the cassettes, sutures or staples are hidden in tissues. The presence of the sutures and staples will apparently remain unknown throughout tissue processing as it does not distrupt the processing. It will usually be found during embedding and most of the time, during shaving. Which is pretty unpleasant. Not only will the staple or sutures cause the blade on the microtome to be instantly blunt, it will also require much digging and pulling out the staples and sutures from the hardened wax block. Fourthly, tissue blocks containing certain breast tissues are not required to be shaved. This is because they are exceptionally thin. Finally, i have to ensure that the wax in the tissue blocks are completely hardened before it can be shaved. This is to prevent the risk of the tissues dropping off from the block. When that happens, the tissue will have to be sent for re-blocking.

I realize that embedding affects shaving a lot. This is because, at times, the tissues are not on the same level in the wax. This is probably due to the tissues not being pressed down towards the mould during embedding. When the tissues are not on the same level, much shaving needs to be done to expose every part of the tissue. When this happens, the risk of losing parts of the tissues increases.


Nurathirah
Tg01
0606561I

11th Week of SIP

Name: Hardina Hamzah

Topic: Microbiology

Title: Processing of Urine Specimens

Urine culture is commonly done to determine whether the patient has urinary tract infection. A urinary tract infection is a bacterial infection that affects any part of the urinary tract. When bacteria get into the bladder or kidney and multiply in the urine, they cause a UTI. Enterobactericeae species such as, E.coli, Kleibsiella pneunomiae, Proteus mirabilis and Pseudomonas aeruginosa, cause most of the urinary tract infection.

Retrieved from: www.pharmacy-and-drugs.com

There are two types of media used and there are TSA with sheep blood agar and MacConkey agar. TSA (Trypticase Soy Agar) with sheep blood agar is useful for the cultivation of a wide variety of fastidious microorganisms. TSA is a general purpose media produced via enzymatic digestion of soybean meal and casein and TSA is frequently the base media of other agar plate type. In this case, it is the base media for blood agar plate. MacConkey agar is a culture medium to grow Gram-negative bacteria and stain them for lactose fermentation. It contain bile salts (to inhibit Gram-positve bacteria), crsytal violet dye (also inhibit Gram-negative bacteria), neutral red dye (to stain microbes fermenting lactose), lactose and peptone.

TSA with Sheep Blood Agar

Retrieved from: www.madsci.org

MacConkey Agar

Retrieved from: www.madsci.org

The urine specimen need to be processed immediately as if prolong, the results may not be accurate. The urine is mixed well before inoculating it onto agar plate. 1ml calibrated disposable loop is used and the loop will be immersed into the urine specimen bottle. The sample will be inoculated onto Blood agar plate first and then MacConkey agar plate by making a straight vertical line down the centre of plate and will be followed by series of close perpendicular streaks through the original line. The plates will then be incubated at 35°C aerobically overnight. At the same time, microscopy cell count of the fresh urine specimen has to be done. The urine sample will be transferred to the notch on the slide chamber of KOVA GLASSTIC SLIDE 10 with grids. The urine sample will be transferred by using the capillary tube and the urine sample will be drawn into the chamber resulting in a homogenous suspension of sediment. The cells to be counted are epithelial cells, white blood cells and red blood cells. When there are a lot of white blood cells (eg. >100 cells) that are counted; this can indicate that the patient may be having urinary tract infection.

KOVA GlASSTIC SLIDE

Retrieved from: http://www.cenmed.com/productDetail.asp?productid=17291&catID=&category=5617&mainCat=&cat=5305

After office hours or during weekends/public holidays or even there is a delay in transporting urine specimen, the dipslide will be used. The dipslide has three types of media which are Cysteine Lactose Electrolyte Deficient (CLED) agar, MacConkey agar and E.coli agar. CLED agar is a valuable non-inhibitory growth medium used in the isolation and differentiation of urinary organisms. Being electrolyte deficient, it prevents the swarming of Proteus species. Lactose fermenters produce yellow colonies on CLED agar; non-lactose fermenters appear blue. It has a pH of approximately 7.3. The staff nurse will dip the dipslide into the container containing patient’s urine and then seal the dipslide into the bottle. The staff in the laboratory will help to incubate overnight. One disadvantage of using this method is that there will be no urine microscopy cell count.

Uricult Trio Dip Slide

Retrieved from: http://www.oriondiagnostica.com/files/odextra/Clinical%20Microbiology%20brochure/782-03GB_Uricult.pdf

The cultures and dipslides which are made on the previous day will be read on the next day. Usually when there is a mixed growth of bateria (different types of organisms growing), it is considered insignificant and will be recorded as negative. However, when there is predominant growth of only one type of bacteria, it is considered as significant and will be recorded as positive. Enterobacteriaceae species can be identified easily by seeing the morphology of the colonies and to confirm the species, indole test can be done. For example, when E.coli is suspected to grow, indole test is done on the spot to confirm. A small filter paper is used and a drop of indole is placed on the filter paper. Then, the filter paper is rubbed against one of the colonies. When the filter paper turns blue, it indicates positive and it is proven that it is an E.coli. Then the culture will be sent for further test which is sensitivity testing. However, when there is a doubt on the identity of the organism, the organism will sent for identification and sensitivity test.

week 10!

Subject title: Clinical Chemistry
Name of test: Urine Microalbumin Qualitative Test


Yay! 10 more weeks to go =)


Principle:

• Urine Microalbumin Qualitative Test is a screening test to determine the presence of albumin and creatinine in urine. An albumin-to-creatinine ratio is also determined since the amount of albumin in the urine and the concentration of urine varies throughout the day, hence if we were to take the albumin results, it would not be very accurate. Creatinine on the other hand is excreted out on a consistent basis and its level is relatively stable in the urine. This test can be considered semi-quantitative, however it is not a confirmatory test.
  

• This test involves plastic strips which contain two reagent areas that test for albumin and creatinine. The strips are then read instrumentally using CLINITEK STATUS®

• Normally, albumin is present in urine at concentrations of less than 20mg/L. Measurements of 20-200mg/L indicates microalbuminuria whereas results of >200mg/L indicate clinical albuminuria. It is abnormal to have protein in your urine as the glomerular basement membrane is actually poorly permeable to it. Microalbuminuria can be an indication of glomerular damage.

• Test principle for albumin: It is based on dye binding using a high affinity sulfonephthalein dye. If albumin is present, the colour of the reagent area that test for albumin will change to blue. The colour ranges from pale green to aqua blue.
  

• Test principle for creatinine: It is based on the peroxidase-like activity of a copper creatinine complex that catalyzes the reaction of diisopropyl-benzene dihydroperoxide and 3,3,'5,5'-tetramethylbenzidine. The colour ranges from orange through green to blue.

Materials:

1. Patients' urine


2. CLINITEK® Microalbumin Reagent Strips

Retrieved August 31,2008, from http://diagnostics.siemens.com/webapp/wcs/stores/servlet/ProductDisplay~q_catalogId~e_-111~a_catTree~e_100001,1015867~a_langId~e_-111~a_productId~e_172993~a_storeId~e_10001.htm

3. CLINITEK® STATUS analyser

Retrieved 31 August, 2008, from http://diagnostics.siemens.com/webapp/wcs/stores/servlet/ProductDisplay?productId=172991&storeId=10001&langId=-111&catalogId=-111&catTree=100001,1015867

Method:

1. Check labels on the urine container to ensure the name of the patient tally

2. Key in the patient's lab no in the analyser

3. Dip a reagent strip into the sample making sure both reagent areas are wet

4. Remove the strip and drag the edge of the strip against the rim of the urine container to remove excess urine

5. Place the reagent strip with the reagent areas facing up, onto the analyser test table. Press START and the table will be automatically pulled into the analyser where the strip will be read.

6. Record the results displayed on the analyser

Results:

• The results are determined by the Albumin-to-Creatinine Ratio (A:C) which is calculated automatically by the analyser
• The results are recorded as NEGATIVE when the A:C displayed is <30mg/g>
• The results are recorded as ABNORMAL when the A:C displayed is 30-300mg/g
• The results are recorded as HIGHLY ABNORMAL when the A:C displayed is >300mg/g

Note:

• Both ABNORMAL and HIGHLY ABNORMAL samples will be confirmed with the quantitative test which can be carried out in the machines COBAS or MODULAR P used in our lab
  

Nur Farhana Binte Ramlan
0604834B

9th week

hello everyone..
this is the 9th week of our SIP..
how's things going?
it has been a busy week for both me and my buddy as we have started embarking on our major project.
we have been dividing our time between benchwork and MP during this time.

for the past weeks, i have been posted to different department which includes urine section, order entry and chemistry section. in urine section, i was introduced to the routine work. basically it the same routine work that has been mentioned by dyana in her latest entry. beside doing urinalysis, the med tech there also have to perform any arterial blood gas (ABG) request. i was only allowed to observe how they process the test as it require the id of the person performing the test. it was rather an easy test to do as they just have to load the syringe into the machine and the machine will produce the result.

i was then attached to order entry section the week after. as i had been there before, i was already familiarise with the work flow there. only that this round i was station to do the order entry itself. there was so many codes to remember. but thankfully there was people around to help me out. there was also files which contain the codes placed near the counter. we can always refer to it. apart from having to know the codes for the different test, we must also know how to labeled the tubes for the different test.
EDTA tube are commonly used for FBC and GHB.
HEPARIN tube can be use for most of the tests. this applies to GEL tube as well as PLAIN tube. FLUORIDE tube are used mainly for glucose, particularly fasting.
if the blood are less than half of the tube, we hav to transfer to aliqoute it out to a secondary tube before performing the test. if the blood is too little, another tube will be use called the hitachi cup.
after labelling, we will then give the tubes to the different sections.

chemistry section was fully automated. we just have to load the specimens into the machine and the machine will produce the result. and as a med tech, we will have to anaylse the results and validate them. however there are certain things that we have to take note when loading the tubes into the machines.
there are 3 machines that are used in chemistry. they are MPA, SWA and COBAS.
MPA is the machine that consists of centrifugation, decapping of the tube's cap and aliquoting the specimen according to the test requested.
in MPA we cannot load citrate, edta and fluoride tube. the tubes in the same rack has to be of the same height. we can load 5 tubes on 1 rack.
SWA and COBAS are those that performed the tests.
there are tests that are only available for COBAS and some that are only available for SWA. we have a list of those tests placed near the chemistry processing table which we can refer to if we are not sure which machine to use. those test that are not listed can be loaded into either one of the machines.
there are times wen the MPA will be fully loaded with samples. so, for urgent cases we usually have to spin the sample manually. we called it 'offline'. whenever we are told spin 'offline', it means that we have to use the secondary tube for the sample.
sometimes, it gets pretty confusing.

i guess that's all i have to share for this entry.
thanks for reading.

sutiana
tg01
0604651j

8th week

for the past 3 weeks, i have been posted to the cytology lab. the atmosphere in the cytology lab is definitely much cosier, probably because the specimens received in the cyto lab is much lesser as compared to the amount of specimens received in the histo lab.

In cyto lab, specimens are categorized into two fields; gynaecological specimens and non-gynaecological specimens. gynaecological specimens are cervico-vaginal smears while non-gynaecological specimens are sputum, urine, CSF and other fluids(peritoneal fluids etc).

1. The gynaecological specimens are received along with the requisition forms.
a) Ensure that the identification number on the slides tally with the number of the forms.
2. The specimens are then fixed in 95% alcohol.
3. The specimens are then loaded into a rack to be placed in the pap stain machine.

Principles of pap stain.
The papanicolaou stain method is a polychrome staining reaction. It is used to portray the variations of cellular maturity and metabolic activities. This method can be used for cervico-vaginal smears and smears from the different bodily secretions which includes respiratory and digestive, to detect the presence of cancer cells. The pap stain involves 3 different types of stains; the Harris haematoxylin, Orange G and EA50. The harris haematoxylin will stain the nuclei blue. EA 50 is made of three elements. The different elements are eosin, Bismarck brown and fast green. The combination of Orange G and EA50 is responsible for the different range of green, blue and pink hues to the cell cytoplasm according to the degree of keratinization of the cell. The cytoplasm of non-keratinized, normal superficial and intermediate squamous cells are stained green. However when the cells are keratinized, the staining becomes orange and pink.

Note: The slide with the specimens smeared in a thin, monolayer manner will give the best staining results. This is because the thickness of the section will affect the intensity of the staining.


4. The stained slides are then placed in a container filled with xylene.

5. The slides are mounted manually.
a) Just enough depex is filled on a cover slip.
b) Take a slide from the rack in the xylene and place it on the cover slip.
c) Ensure that there are no bubbles present between the slide and slip.
d) If bubbles are seen, press the slip gently to allow the bubbles to escape.
e) If there are too many bubbles seen, the slide should be remounted.
f) To remount the slide: Place slide in a container with xylene until the cover slip dissociates
and until the mounting medium is removed. Then, mount as usual.

6. The mounted slides are left to dry.
7. Label the slides according to the lab number.
8. The slides are ready to be screened.


Nurathirah
0606561I
Tg 01

7th week of SIP

Topic: Haemotology

Hey. I've been attached to Haematology lab for 3 weeks and i think, it's the most busiest lab. In this post, i'm going to share about Bleeding Time and Dengue Serology.


Bleeding Time


Bleeding time is done to estimate the integrity of haemostatic plug which will be formed at the site of injury to arrest bleeding. It is also used in the diagnosis of haemostatic defects and platelet dysfunction. A minor incision is made on the forearm and the time between the infliction of incision and the moment bleeding stops is measured. The formation of haemostatic plug on the site of injury depends on the adequate amount of platelets to adhere to the subendothelium and to form aggregates. A prolonged bleeding time does not itself diagnose underlying platelet disorder. It does indicate the need for further tests such as platelet count, platelet aggregation studies etc.

The items needed to conduct the test for bleeding time are sphygmomanometer, filter paper, antiseptic swab, stopwatch, plaster and single-use Surgicutt blade. The sphygmomanometer cuff is wrapped on the upper arm. Cuff will be inflated to 40mmHg and will remain at exact pressure throughout the test. The time between inflation of cuff and incision should be 20-60seconds. For a newborn, the pressure should be maintained in accordance with the weight of newborn. (<1kg-> 2 kg- 30mmHg) The forearm must be cleaned with alcohol swab before incision. Area with veins, scars or bruises should be avoided. Minimal pressure is used on Surgicutt device so that it lightly touches the skin. After incision, the timing will start. Every 30seconds, the blood is blot with the filter paper until it no longer stains the paper. For neonates lesser than 5 months, the test can be stopped at 5 minutes and bleeding is reported as more than 5 minutes. For adult and pediatrics, the test can be stopped at 20 minutes and bleeding time is reported as more than 20 minutes. The reference range based on Surgicutt’s insert, 2 to 8 minutes for adult, 1.30 to 8.99 minutes for junior (5 months to 15 years old) and 0.85 to 1.65 minutes for newborn (1 to 4 months).

The estimation of the integrity of haemostatic plug measures the interaction between the capillaries and platelets, platelet defects and vascular disorders. It measures the interaction between the capillaries and clotting mechanisms in vivo. However, the test does have limitations. It is not always positive in platelet function defects because of the mildness of the defect or variability in the condition. The results of this test alone are not sufficient to diagnose specific conditions. Further tests are needed.


Dengue Serology
Dengue serology test is performed to obtain a qualitative presumptive detection of IgG and IgM Dengue, a flavivirus, is found in large areas of the tropics and subtropics. Transmission is by mosquito, principally Aedes aegypti and Aedes albopictus. Dengue virus infection causes a spectrum of clinical manifestations ranging from asymptotic to fatal haemorrhagic disease.


When the virus is present in the patient's serum, dengue-specified IgM or IgG antibodies bind to anti-human IgM or IgG antibodies immobolized in 2 lines across the cassette membrane. Colloidal gold complexes containing recombinant dengue 1-4 antigens are captured by the bound patient's IgM or IgG to visible pink line(s).


Procedure:


1. Centrifuge the blood sample at 3000rpm for 10 minutes.


2. Sample serum should be preferably clear, non-haemolysed, non-lipaemic and non-icteric.


3. Before running the assay, allow the buffer to equilibrate to room temperature after taking it out from the fridge.


4. Remove cassette from pouch.


5. Add 10uL of serum in circular well using mechanical pipette.


6. Allow sample to absorb entirely into specimen pad within circular well.


7. Add 2 drops of buffer to square well at base of cassette. Ensure buffer bottle is held vertically.


8. Read results in exactly 15 minutes after adding buffer. All reactions are visualised as pink lines.


9. For assay to be valid, the control line must appear.

Retrieved from: http://www.panbio.com.au/download/Dengue%20Duo%20Cassette%20procedure.pdf

The picture is not very clear. So, just go to the website and see.

If only the control line appears, the test is regard negative. However, if dengue infection is strongly suspected, the doctor may consider to conduct PCR for dengue virus within 5 days.

If control and IgM lines appear, it is a primary dengue infection.

If control and IgG lines appear, it is a secondary dengue infection.

If all lines appear (control, IgM and IgG), it is a secondary infection.

Control line must appear everytime. If it does not appear, the test is regard as invalid.

Primary infection is characterised by the presence of detectable IgM antibodies 3 to 5 days after onset of infection. Secondary infection is characterised by the elevation of specific IgG antibodies 1 to 2 days after onset of infection and in majority of the cases, this is generally accompanied by an elevation of IgM antibodies.

Hope u guys understand.

Hardina Hamzah
Tg01

6th week of SIP

Hi everyone! For the last 2weeks, I was attached to the Urinalysis/Routine section. Well, it was interesting to look at the different colour of urine specimen. Do you know that urine are not always colourless yellow?Some are bloody yellow, and some are dark yellow and are very turbid. Yucks. Haha, but as the 2nd weeks pass by, it became normal thing. In this section, I get to observe and practice on the different test done on urine sample. Some of the tests done at the lab are Urine FEME (Full Examination/Microscopic Examination), Blood Ketone Test, Urine Pregnancy Test, Feccal Occult Blood Test, Ove, Amoeba and Cyst Test and etc..


Topic: Urine FEME (Full Examination/Microscopic Examination)

Urine FEME are microscopic examination of urine sediment. It is one of the common test done in the lab. Everyday, there will be lots of requests for this test. Urine specimen is examined unstained and used to access for sediments such as red blood cells, epithelial cells, white blood cells, casts, crystals, microoragnisms, blastoconidia, parasites and spermatoza. Random urine sample received from the processing section are mixed well by inverting the container about 5-8times. The urine is then transfered to the notch on the KOVA Glasstic Slide Chamber with a plastic pipette. The urine(about 6.6ul) will be drawn into the chamber by the caillary action. According to the SOP, to examine and quantitate casts and epithelial cells, use low power magnification which is 10X objective. While to quantitate all the other cells, use high power magnification which is 40X objective. Usually the MedTech would view the grid using the 20X objective to evaluate whether it is a low cell count or high cell count. They would then proceed to count either 10 small grids within different quadrant if it is a high cell count or count 4 complete quadrants of the grids if it is a low cell count. Cells/ul are calculated by multiplying the average cells obtained per grid by 90.




KOVA Glasstic Slide Chamber

Picture taken from:
http://www.cardinal.com/mps/catalog/ASP/U3050-11.asp?cat=Laboratory




This is how the grid would look like.
If it is a low cell count (few cells), only the four quadrants are count.





If it is a high count, 10 smaller grids are counted.


All urine sample which request for Urine FEME will be run in the Clinitek Atlas Analyser. It is a fully automated reflectance spectrophotometer which uses Clinitek Atlas Reagent Paks for testing glucose, bilirubin, ketone, occult blood, pH, protein, urobilinogen, nitrite, leukocytes, and the colour of the specimen. The analyser also helps to determine the specimen's specific gravity using the refractive index method and the clarity by measuring the transmission and scattering of light that passes through the specimen. Basically, the Clinitek Atlas Analyser acts as an automated dipstick. However, if the analyser is down, the manual dipstick test have to be done. Last week, I experience a situation in which the analyser was faulty and we had to the the manual dipstick method. It was hectic as there were so many urine samples and we had to read the result one by one and record it down before entering it into the LIS. Fortunately, the engineer came to the rescue and fixed the analyser.



Clinitek Atlas
Picture taken from :
http://diagnostics.siemens.com/webapp/wcs/stores/servlet/ProductDisplay~q_catalogId~e_-111~a_catTree~e_100001,1015867~a_langId~e_-111~a_productId~e_172988~a_storeId~e_10001.htm



Topic 2: Urine/Serum hcG Pregnancy Test




hcG combo pack

Urine pregnacy test is one of the common and simple test. It only takes 5 miunutes. The test uses the TestPack hCG Combo which is a rapid immunoassay for qualitative detection of Human Chronic Gonadotropin (hcG) in serum and urine for early detection. hcG is a glycoprotein hormone that is produced by the blastocyst. The specimen used in this test is a random urine specimen collected in a clean and dry container. It will be best if collection is done on first morning specimen as it contains the highest amount of hcG. Urine sample is sent to the Urinalysis section. Urine is drawn to the line marked in the transfer pipette. The entire content in the pipette is drop into the sample well on the reaction disc. Sample is allow to react with the test kit.

In the test procedure, urine or serum is added to the sample well of the reaction disc with the aid of the transfer pipette and is allowed to migrate through the membrane. The urine or serum will pass through the membrane and thereby mobilizing the anti-hcG monoclonal antibody-colloid. The antibody-colloid complex migrates through the membrane and is captured by the immobilized anti-hcG polyclonal antibodies in the result window, providing a visual indication of the presence of hcG. Since the test only takes 5 minutes, reading should be after the 5 minutes.Any result that appears after 5 minutes are to be ignored. If hcG is present in the urine or serum at levels of 25mlu/ml or greater, a plus sign (+) will appear in result window. If no hcG is detected, a minus(-) will appear. Thus. providing an easy intepretion of result for patient's sample.

Pre-menopausal females generally contain < 5mlu/ml hcG while healthy males and post menopausal females generally contain < 10 mlu/ml. In pregnant women the hcG level rises within 9-12days after ovulation and the urine hcG reaches level of up to 1500mlu/ml 8-10 weeks after last menstrual period.

Thank You for reading!

Peace,
Dyana

5th week of SIP

hello! how's everyone? hope you guys are doing great. coz i'm pretty much enjoying my attachment here( somewhere)..hehe
well it has been 5 weeks since we started our SIP..how time flies..
for the past 5 weeks, i was posted to Microbiology department as well as Haematology, micro being my longest stay for 3 1/2 weeks..
so im going to focus on micro for this entry..

the micro department here is pretty much on its own. it has a place by itself unlike the rest of the department. it is always these same people that will be runnig the tests for the patient. they are like our specialist team. unlike the rest of the department, micro is manually run. they are not automated except for blood culture where we use a machine called BATEC. i will talk about it more as in the later part. basically, there are three sections in micro- processing, reading for urine culture and reading for miscellaneous culture.

in micro, we process the order entry on our own. before entering the patient's demographic information, we will make sure that the name on the request form matches the one labeled on the specimen. then we will enter into the LISS. after which we will do the requested tests. there are many tests that is performed by the medical technologist. hence i will only mentioned a few test here( if i mentioned every single test, i believe you people will get bored reading it). so one of the test is urine culture. urine culture require two agar plates- TSA blood and Cled. we divide the TSA plate into 4 and the Cled into 2. the streaking for the urine test is as follows:

streaking on Cled plate

streaking on TSA blood

the next test is MRSA. for MRSA, we only use one plate which is mannitol. we divide the plate into three and the streaking is a follow:

streaking on Mannitol plate

we also do wound culture. wound culture requires 4 plates- TSA blood, Scandler blood, MacConkey and PEA. the streaking is the same as on mannitol.

apart from the test that requires culturing of specimen, here in micro we also performed test using kits. one of the test that uses kit is the test for S.pneumonia and Legionella in urine specimen. the test is very simple to performed.
1) we use the cotton bud and dip into the urine sample.
2)we place the cotton bud in the kit test.
3)we place three drops of reagent into the hole provided, two drops for Legionella (different reagent for different test)
4)we leave it for 15 mins before reading the kit
5)if only the control line appears, it is negative, we both the control line and the positive line appears means its positive

the next test kit is for the detection of Clostridium difficile toxin A and B.
the test is not that easy as compared to the test earlier. basically there are 4 reagent use in this test- dilute, conjugate, wash buffer and substrate.
1)first we put 500ul of dilute and 1 drop of conjugate into a test tube.
2)then we take about 5mg of stool and mix it with the two reagent. ( there must be no clump of stool presence in the mixture)
3) pipette about 1ml of the mixture into the test kit(the small hole) and wait for 10 minutes
4) when 15 mins is up, pipette 300ul of wash buffer into the kit (the big hole) as well as 2 drops of substrate
5)leave it for 10 mins before reading the result
6) the interpretation of the result is the same as the test mentioned earlier.

*i know pictures will help. but i cant find one on the net that resemble the test kit that i mentioned. i will try to get the pics and post it here ASAP. ok..sorry

as promise, i will now talk about the blood culture process. the blood will come in 'blood bottles',
one has a blue cap and the other is gold cap. it is recommended to send the blood in both bottles. if only one bottle is sent, we can still run the test. so after entering the information into LISS, we will incubate the 'blood bottles' in Batec. Batec is a specialized machine for blood cultures. we have to incubate the 'blood bottles' for 5 days. no bacteria grow after 5 days will indicate a negative results. for positive blood, it will shows the results anytime between the 5 days. if the blood is positive, we will then have to culture them on the appropriate agar plates.

blood bottles


all the cultures process will have to be incubated overnight. reading will only be done the next day.

reading of the plates includes identifying the bacteria that is present. plates where colonies are seen (more than 20 colonies) will have to be followed up with various test for identification purpose.
example of the tests are catalyse,oxidase,staph-plus,indole and many more.
each test have its own specificity for identifying the bacteria.


sutiana
0604651J
TG01

4th week comment-replies

due to the generous amount of responses, i have decided to counter each of them in a post.

THE CODEC 5 said...

Hi,

So far in your attachment, have you met any difficulties? If there is, can you share?

By the way, you mention that if the orientation of the tissue is not done properly, then it will hinder the sectioning. So what happen if this really happens? Does your lab redo it or just leave it as it is? The samples obtained are limited right?

Thanks. That's all.

Xin Yi
TG02

xin yi-
so far, i believe that it has been a pretty smooth-sailing ride on my part. nothing lethal whatsoever. haha. though initially, i had a minor issue on adapting to the working environment and understanding the interpersonal dynamics. but time has truly rectify my concerns.

well, if the large tissue is not properly orientated(meaning not of the same level), the tissue block has to be shaved deeper so that a full face can be achieved. however, if the thinner tissues are not embedded to the paraffin wax at the same level, they will have to be re-blocked. this is because shaving the thinner tissues more will risk losing the patient's tissue.

~immortals~ said...

olla!!!!
aint it abit gruesome to see all those parts of the body??

anyway, yew saed that orientation of the tissues are crucial, such as the orientation of the cyst.

so my question is, why is the cyst wall the main interest?

and why must the cyst wall be at the base of the mold, and not be anywhere else?

mayafirhana
tg02

maya-
haha initially, it put me off a bit. but after a while, i am pretty much accustomed to it. A cyst is an abnormal, sac-like structure within a tissue that can contain fluid, air, or semisolid substance. it can be present at anywhere in the body and varies in size. The outer portion of a cyst is termed the cyst wall. i believe that the cyst wall is the most crucial part to produce a diagnosis. the cells at the wall will show if it is benign or malignant. oh, it has to be at the base of the mold because if you recall embedding right, you will always section the surface of the tissue which is at the base of the mold. do you recall? hehe =)

~immortals~ said...

Hi Tira!!

U mention that the specimens for surgery are sent to the specimen recieving room. How long are they usually kept there before being processed?

Another quest: What are the possible errors that might arise in the embedding process?

opps my name is AmiR!!
TG02


Amir-
hello Amir!! Specimens FROM surgery are sent to the receiving room to be correlated with the forms. subsequently, they are sent to the trimming room. depending on their sizes, they are either trimmed by the pathologists or immediately placed into the cassettes and fixed in formalin. and after that, they are placed in the processor. so the duration from being in the receiving room to being processed varies. larger tissues, longer duration. hmm errors during embedding..when the paraffin wax has not completely cooled down and it is immediately being sectioned, the tissue will break. that's when re-blocking has to be done.

tg01 group 2 said...

Hi Athirah,

Some questions to ask you,

1)You mentioned that the forcep is hot and I understand it is sterile.
Wouldn't the heat from the forcep breaks the tissue?

2)What do you use to press the tissue flat down on the mold and how do you ensure this is done properly?

Thankz!

Han Yang
TG01

hey han yang,
1) oh no. it would not. the temperature of the forceps is about 65-70 degrees celsius. At this temperature, the tissue will not break. The heated tips of the forceps actually helps to prevent the tissue from sticking to the tip and hence prevents carry over which could lead to contamination.

2)we use forceps and a flat-surfaced tip made of pure metal.

BMT said...

Athirah!
Ok I'll bug you with a question. Which I'm sure will be arguably easy for you to answer since you seem to be a pro at Histo already. =P

Anyway, must all tissue samples (like you have mentioned the cyst) be placed at the base, or can they be placed in the center?

Elyana
0606676E
TG01

dear elyn-
it has to be placed at the base so that sectioning can be done efficiently. imagine if it is placed at the center of the mold, the process of sectioning will be prolonged.

~immortals~ said...

miss nurathirah,

You mentioned the use of an electrically heated forceps for the transfer of the tissues into the mold.

I would like to know at which temperature are the forceps heated to, as i believe that a temperature which is too high may damage the tissues.

I would greatly appreciate if you could enlighten me on this matter.

thanks!!!!

fellow med tech,
rusydiana

Rusyyy-
the temperature of the forceps is about 65-70 degree celsius. so far i have not witnessed any tissue damage at this temperature. also the electronic forceps have overheat protection, to ensure that the temperature of the forceps do not go beyond 70 degree celsius.

BMT said...

Hi:) can i ask u if the machine used to fill the wax is the same as the one we have in our sch lab?thanks:) because i am curious how does it fills the wax:)

Rachael
Tg01

hello there rachael-
honestly, i can't remember exactly how the embedding machine in school looks like. haha. well since there are many types of embedding machine. however, i can assure you they consist of the same parts; the wax dispenser, the cold plate, the hot surface, the mold storage area.


i'd appreciate it if you'll correct me if i'm wrong. thanks.

Nurathirah
Tg01