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