Friday, August 24, 2007

Student Internship Programme (SIP) HAEM

Are you surprised that haematology department also performs urinalysis? I hope you do, because I, for one, certainly did. The difference between the urine examinations in various departments lies mainly with the type of tool used for analysis. The principles are similar in that common findings such as RBCs, WBCs, epithelial cells (EC) are counted and reported. Kova glasstic 10 has 10 chambers[able to charge 10 different urine samples] and in each of the chamber, there lies 81 small grids.

Name of Test: Urine microscope examination using Kova glasstic 10 with grid slide

Principle of Test: Undiluted and uncentrifuged urine is drawn into the Kova slide 10 chamber for microscopic examination and identification
Type of Specimen: "Early morning" or random urine, midstream clean catch samples
[Morning-voided sample preferred as it is more concentrated than random samples during the day as urine could be diluted due to increased fluid consumption; giving false picture of a patient's health]


1. Check that name on form tally with sample.

2. Assign a lab number and stamp for WBC, RBC, EC grid on request form.

3. Mix urine thoroughly by swirling several times before opening.

4. Record the appearance(colour/turbidity) of sample.

5. Charge 9µl urine on the Kova glasstic slide chamber.

6. By capillary action, 6.6µl of the urine will be drawn into the chamber resulting in a homogenous suspension of the sediment. (you may ask how, but there is a tiny compartment under the chamber where excess urine goes and only the stipulated amount of urine will be held in the chamber)

7. Allow 10 minutes for the cells to settle. (as urine continues to flow within the chamber)

8. Examine for urinary sediments and quantitate casts/crystals at low power 100x

9. Quantitate all cells at high power 400x. Count the cells within the lines of the small 0.33mm square grid.
a) For low cell count sample: count the total cells of each specific type contained in 36 small grids of the counting grid. Write the values beside the stamp 'WBC', 'RBC' and EC accordingly and write 36 beside the stamp grid.
b) For high cell count sample: count the total cells of each specific type contained in 10 small grids of the counting grid. Write the values beside the stamp 'WBC', 'RBC' and EC accordingly and write 10 beside the stamp grid.
c) For very high cell count sample: count the total cells of each specific type contained in 1 small grid. Write the values beside the stamp 'WBC', 'RBC' and EC accordingly and write 1 beside the stamp grid.

10. Enter the reported values and the LIS system will compute the resules and convert the values to per µl.

11. Transfer the computed values to the request form on the appropriate space.

Reference ranges:
WBC: 0-6/ µl
RBC: 0-3/ µl

-Alex Tan Tg02 0503222B
To dorothy

Flow cytometry is a technology that stimulataneously measures and then analyses multiple physical characteristics of single particles, usually cells as they flow in a fluid stream through a beam of light. The properties include the particle's size, granularity and fluorescence intensity. samples are incubated with monoclonal antibodies conjugated with flurochrome which are excited by laser.Physical properties are measured as the flurochrome tagged cells pass through the laser light.

reference range of CD4 : 28.2-50.7%
reference range of CD8: 12.5-38.5

To kangting

here's the ans

reference range if CD4 : 28.2-50.7
reference range of CD8:12.5-38.5

To phuiyuen

This test can be performed after renal transplant to see whether is there any sign of rejection by the recipient. CD4 level should be dropping and CD8 level on the rise.

To zahirah

CD4 T-cells are a type of lymphocyte (which is one type of white blood cell). They initiate the immune system’s response to viruses such as HIV. CD4 cells can be infected and killed by HIV, so the CD4 count generally declines. CD8 T-cells, on the other hand, are the immune system’s killers of abnormal or infected body cells. As HIV infection worsens, the number of CD8 cells present in the body tends to increase.

Flurochorme labeled antibodies are used such as FITC and PE.

To jiahao

Ya. it is also possible for leukaemia.

Take b-cell type Chronic lymphocytic leukaemia as an example, the cell markers are CD5, CD20,CD52,CD23,CD19 and CD37.

To Michelle

In this case, i am talking about CD4 and CD8, these two surface markers. CD4 is for T- helper cells and CD8 for T-supressor cells which both have their respectively specific functions which i mentioned in my post. As for the subgroups, CD4 and CD8 are actually subgroups of mature T lymphocytes and the surface marker for it is CD3. CD4 and CD8 are subsets of CD3.

To Yvonne

well, this test is not a confirmatory test for HIV. other tests are needed to carry out to confirm the diagnosis.

Saturday, August 18, 2007


section posted: flow cytometry lab

Name of test: CD 4 and CD 8 count

Purpose of the test : it is used as a diagnostic and prognostic tool for clinical classification of immunodeficiency states such as autoimmune state and immune monitoring of responses to therapy such as to monitor progression of AIDS by measuring CD4 and CD 8 levels.
CD4 is expressed by T-Helper cells (collaborate with B cells promoting their proliferation and promote t cytotoxic cells development.)
CD8 is expressed by T- suppressor cells (suppress immune responses)

Principle of the test: flow cytometry has been commonly used for the measurement of surface marker antigens by fluorescent labeled monoclonal antibody which will pass through laser beam generating scattered and fluorescent signal that can be processed by computers. These surface makers are related to specific functions and subgroups.

Clinical significance: CD4 and CD 8 measurement are used to monitor progression of AIDS as CD4 cells are depleted by HIV infection whereas CD8 cells persist.

well, that's all! have fun for sip..

kai lin

Sunday, August 12, 2007

Rountine Laboratory

I was posted to the routine laboratory which deal with all body fluid, these include urine, peritoneal fluid, CSF and semen as well. Today i will be posting about semen analysis which is a pretty interesting topic itself.

Semen analysis is the testing for male infertility problems. This test provide information about the quality and quantity of the sperms. The semen sample is analysed for volume, viscosity, pH and colour of the ejaculate, sperm concentration, motility and morphology. It is also examine for presence of RBC or WB which is often an indicator of infection, inflammation.

The ejaculate should be obtained after at least 3 days of abstinence from sexual intercourse and is best produced by masturbation. The entire ejaculater is collected in a clean sterile container. Condom must NOT be used as it is spermicidal. The sample must be examined within 1-2 hours after collection.

a. Measure the volume received
b. Note the appearance; record and abnormalities such as change in colour, decreased in turbidity or presence of blood.
c. Note the change in viscosity.
d. SQA 11 C- P will provide information on progressive motility and normal morphology
e. Aggluntination- exmaine under microscope for clumping of spermatoza
f. viability:
1) Add a drop of seminal fluid to 1 drop of eosin stain in a 75x 10 mm test tube
2) Mix and stand for 5 minutes
3) Examine under 40 x objective for viable spermatoza. Dead spermatoza will be stained pink
g. pH - Dip a pH indicator strip into the seminal fluid and record the result using the standarh chart
h. Cell count (density)
1) Mix the seminal fluid thoroughtly and dilute fluid to a 1:20 concentration
2) Mix and count in KOVA slide
i. % motility -Mix specimen well. Place a drop into the glass slide and read microscopically. ( Note the quality of motility and progression)

a. Volume: norma volume ranges between 1.5-6.0ml
b. Appearance:
1) Colour- normal semen appear grey to white opalescent fluid. Different colour may indicate problem
2) Viscosity - hyper viscosity may impair sperm motility
3) Liquefaction - Normal time frame is 10-30 minutes.
c. Normal sperm count is > 20 x 10^6sperm/ml
d. Motility - sperm motility study identifies the number if motile sperm seen in an ejaculate specimen.
e. Morphology - Nomal sperm have oval head shapes, an intact central and an uncoiled single tail.
f. Aggluntination - presence of aggluntination can be caused by antisperm antibody.
g. pH - normal pH should range from 7.2-8.0

Ching Wei

Sunday, August 5, 2007

SIP online sharing

Hihi to all. Sorry for the late posting as I couldnt log in due to the cookies. It is the 6th week of SIP, time really fly. For the past 6 weeks I been went to Processing, Biochemistry and Immunoassy Section. For the biochemistry and immunoassay section, all of the tests is done by analyser. While processing is understandable is to order entry

For this sharing, I will talk more about immunoassay section. I will be sharing some guideline on HIV screening & confirmation tests and BDvacutainer uses.

In the immunoassay section, the analyser use is the Architect ci2000. This machines uses the chemiluminescent microparticle immunoassay technololgy to determine the presence of Ag, Ab and analytes in the sample.

The BDVacutainer System

Cap Colour: Red
Effects on Specimen: Blood clots, and the serum is separated by centrifugation
Uses: Chemistries, Immunology and Serology, Blood Bank (Crossmatch)

Cap Colour: Gold
Additives: Separating gel and clot activator
Effects on Specimen: Serum separator tube (SST) contains a gel at the bottom to separate blood from serum on centrifugation
Uses: Serology, endocrine, immunology, including HIV

Cap Colour: Light green
Plasma Separating Tube (Na Heparin)
Effects on Specimen: Anticoagulants with lithium heparin; Plasma is separated with PST gel at the bottom of the tube
Uses: Chemistries

Cap Colour: Lavender/Purple
Additives: EDTA (liquid form)
Effects on Specimen: Forms calcium salts to remove calcium to prevent clotting and platelet clumping
Uses: Hematology

Cap Colour: Light blue
Sodium citrate (Na Citrate)
Effects on Specimen: Forms calcium salts to remove calcium
Uses: Coagulation tests (PT, PTT)

Cap Colour: Dark green
Sodium heparin or lithium heparin
Effects on Specimen: Inactivates thrombin and thromboplastin
Uses: Ammonia, lactate, HLA typing

Cap Colour: Dark Blue
Sodium heparin
Effects on Specimen: Forms calcium salts. Tube is designed to contain no contaminating metals
Uses: Toxicology and trace element testing (zinc, copper, lead, mercury) and drug level testing

Cap Colour: Light Gray
Sodium fluoride and potassium oxalate
Effects on Specimen: Antiglycolytic agent preserves glucose
Uses: For lithium level, use sodium heparin. Glucose test.

Guideline to interpretation of result of HIV Screening and Western blot confirmation tests
HIV Screen: Non-reactive
Interpretation: A non-reactive HIV Ab screening test does not necessary exclude the possibility of infection with HIV. If exposure to HIV is suspected and thus serum is taken less than 3 months after this exposure, this should be retested after that time.
HIV Screen: Reactive
HIV Ab Confirmation: Negative
Interpretation: This result is commonly due to the presence of non-specific Ab, but occasionally may be seen in very early HIV infection. If exposure to HIV is suspected, 10ml blood in EDTA is send in 2-4 weeks time for repeat screening. For female patients in late pregnancy, the repeat sample is sent immediately.
(Rationale: HIV Ag is detected 1 to 2 weeks earlier than HIV Ab, thus making HIV screening test more sensitive than the western blot in the window period)

HIV Screen: Reactive
HIV Ab Confirmation: Indeterminate
Interpretation: This reactivity pattern commonly occurs in uninfected individuals due to the presence of non-specific Ab but may also be observed in early HIV infection. If exposure to HIV is suspected, 10ml blood in EDTA is send in 2-4 weeks time for repeat screening. Otherwise, another serum sample taken 6 weekslater will permit diagnosis in the majority cases.
Note: If a second sample is taken after 6 weeks and the western blot remain unchanged(still indeterminate), the following will be the interpretation applies:
- This indeterminate western blot profile has remain unchanged over 6 weeks. This may indicate a negative HIV Ab status associated with the lack of clinical signs and symptoms and/or supported by a negative history of exposure. If repeated exposure is suspected after the first sample is collected, send another sample for repeating screening.

HIV Screening: Reactive
HIV Ab Confirmation: Inconclusive
Interpretation: This is an indeterminate profile that include Ab to envelope and/or polymerase genes, which one often more sensitive indicators of earlu seroconversion. This profile may however also be due to non-specific Ab. 10ml blood in EDTA is send in 2-4weeks time to resolve the result.

HIV Screening: Reactive
HIV Ab Confirmation: Positive
Interpretation: Patient is consider HIV Ab positive

To confirm whether HIV positive or negative take a long procedure. And HIV Ab confirmation is negative does not mean negative too. As to test for HIV Ab confirmation take a period of time after exposure.

Feel free to ask qn but pls be mercy.

Juexiu Tg02