Sunday, 30 September 2007

Specimen Reception and Immunology

Hi all,

Sorry for the late posting.
I have just finished Specimen Reception department in my lab and now currently at Immunology.

At the specimen reception department, it is basically about SOPs and following strictly to SOPs. As we have major clients apart from normal clinics, we have to know our clients well because the specimens received from each client are processed differently. So basically, I spent a long time just learning the procedures because we are expected to work like the other staff there.

It is also essential to know the different types of tubes and their functions because we have to inform the doctors and reject the specimen if wrong specimen types are sent for testing. (It has happened quite frequently)

Long Plain tubes:

Known as serum separator tubes when gel is added to the tube. It contains a clot activator (I don't know the exact ingredient) pre-coated in the tube. When spun down the red cells are at the bottom and the gel in the middle with the serum at the top. It is important to note that different type of gels from different materials may cause inaccuracies. For those without the gel, it only has the clot activator. Long plain tubes are usually used for serum analysis such as VDRL testing.

Heparin tubes:

Usually available as sodium, litium or ammonium salt pre-coated in the tube, it is used for trace elements screening in industrial toxicology for patients working in heavily polluting industries. It is also recommended that for potassium analysis, heparin tubes are to be used. When using the long plain, potassium would be released from the platets giving an increased in serum potassium. EDTA tubes comes with potassium salts and would cause falsely high potassium results.

Fluoride/oxalate tubes:

Oxalate will cause osmotic redistribution between the the plasma and erythrocytes, diluting the plasma as a result (water moves from erythrocytes to the plasma ). This would decrease hematocrit levels, disrupt the morphology and cell membranes of erythrocytes. When the samples arrive in our lab, "water condensation" may be observed inside the tube because of the osmotic redistribution. Therefore, fluoride/oxalate tubes are only used for ABO and antibody screening in our lab. Oxalate will also bind to calcium in the blood, causing a falsely low blood calcium level.

Citrate tubes:
Usually for coagulation studies because it preserves the coagulation factors. Citrate also causes osomotic redistribution and decreases the concentration of many analytes (like fluoride/oxalate tubes). However in our lab, citrate tubes are most commonly sent along with a EDTA for full blood count or as a follow up of an EDTA specimen usually due to possible/observed platelet clumping in certain patients (cause unknown) when the specimen is sent in EDTA.


During my term in Immunology, the common tests run in the department are:

1. Dengue (most common but low sample volume compared to HIV and it is the only manual test.)

Run by the Centaur machine

2. HIV (most common)
3. Tumor markers ( Ca 199, Ca 125 etc)
4. Ferritin
5. Vitamin B 12 and folate acid
6. Hormones such as cortisol and testosterone

Principles of Dengue is the same as Sally's post.
Principles of HIV is the same as Lizzie's post a number of weeks ago.

As promised, I managed to find out about the alternative system to HIV testing with a much shorter testing time. As it is newly tried out in the Serology department, I don't know much about it yet. It is the Roche Modulla from Roche diagnostics.

Ca 15-3

Ca 15-3 is usually for testing of metastatic breast cancer. When a women is diagnosed with breast cancer, usually, her specimen has the presence of cancer-related antigens especially Ca 15-3. Note that for all tumor marker tests conducted, an abnormal result does not mean that ther patient has cancer. There are also conditions where abnormalities are observed in normal healthy patients. Therefore, a confirmatory test is necessary to detect cancer in patients.
Ca 15-3 can be used to monitor response to treatment of disease. In patient with known metastasis, a reduction may mean a good sign of positive response to treatment while an increase may mean the disease progressing to the next stage.

Principle of the test:
2 step sandwhich ELIZA using direct chemilluminescent.

The system will perform:

1. Dispense sample into cuvette

2. Dispense conjugate reagent and solid phase and incubate at 37 deg celcius.
The conjugate is made of monoclonal mouse antibody 115D8 labelled with fluorescein, which is specific for CA 15-3 antigens. The solid phase is made of monoclonal mouse capture antibody coupled to paramagnetic (refer to PIPC 2 notes for definition of paramagnetic) particles.

3. After incubation, the complex (conjugate and solid phase) is washed to remove unbound antibodies.

4. The complex is resuspended using the wash solution and Lite reagent is dispensed. The Lite reagent consists of monoclonal mouse antibody, DF3, specific for CA 15-3 labeled with acridinium ester.

5. Cuvette is washed to remove unbound Lite reagent.

6. Acid and base reagent is dispensed to initiate the chemiluminescent reaction.

High amount of CA 15-3 in patient's sample = high amount of relative light units (RLU) detected. This means that with an increase in CA 15-3, the intensity of chemiluminescence increases.

Once again, I appologise for posting so late.

With regards,
Yeng Ting


Debra said...


May i know what kind of samples are taken/needed for the Ca 15-3 test for breast cancer? Needle aspiration or biopsy or etc?

Also, are there any other outdie factors that can influence the ELIZA reading significantly? If so, what steps are taken to reduce it's incidence? Because, this would influence the results and it'll be hard to give a diagnosis/prognosis no?

- Debra, TG02

first6weeks said...

I alway have a qn regarding tubes so i guess it a good opportunity to ask u, when a patient nd to draw blood according to all respective tubes,in which sequence does he go first or it doesnt make a difference? Eg, citrate followed by EDTA etc

Ching Wei

MedBankers said...


hmmm... my lab uses EDTA-tube for ABO grouping/Ab Screening? or oxalate is EDTA? if so, doesnt oxalate tube more costly than EDTA tube?

MedBankers said...

elaine by the way.. heehee

Anonymous said...

To elaine:

I mentioned that fluoride tubes can only be used for ABO testing. It is recommended to send in EDTA tubes and in my lab, we don't really receive much fluoride tubes for ABO. And to rectify my mistake, the Fluoride tubes are unsuitable for any form of haematological testing(except ABO and antibody screening)and studies such as blood calcium levels which are affected by the osmotic changes. However, studies such as lactate studies and blood glucose (blood glucose testing and OGTT)uses the fluoride tubes for testing.

To Ching Wei:

IN general, there is an order of draw. Usually, phlembotomists would draw blood for blood culture first, followed by citrate, plain, Heparin, EDTA then fluoride. For my lab, the most common tubes are the plain, EDTA and Fluoride. Therefore, I noticed that our plembotomists usually draw in the order of plain, EDTA and Fluoride. There are circumstances where the plembotomist actually so called cheated. THey draw the blood in one long plain and then pour out into the EDTA and Fluoride before the blood clots. Today, I happpen to observe a patient with difficult vein. After 4 tries to draw blood (maximun is 5 tries due to MOH regulations), we managed to get slightly less then a quarter of a long plain tube. THus, the plembotomist poured a little of the blood into an EDTA and an even lesser amount into the Fluoride tube.
Since there is the presence of the protocol for the order of blood drawing, I guess that there is a reason behind the order. For that, I will get back to you ASAP.

Have a nice day,

Yeng TIng

Anonymous said...

To Debra,

For CA 15-3, we use a serum sample taken from in a long plain tube. Needle aspiration and biopsy are the gold standard for the diagnosis of CA 15-3. HOwever in my lab, the objective of the test is mainly for general screening and especially for monitoring of disease progression and response to treatment in known patients with metastasis.

Besides, needle aspirations and biopsies are done in a hospital setting instead of a private setting. Therefore, we don't receive such specimens here.

In my lab, we dump the tubes into the labcell (our total lab automation system) after centrifuging and we don't get to touch it at all. The labcell allocates to the appropriate machines, tells the machine which test to run and keeps the tubes after testing into proper slots in the racks and we keep those racks in the cold room for storage. Results are also automatically uploaded into the LIS. So, the chances of analytical outdie factors are greatly reduced. Factors such as machine faults and pre-analytical factors are not much we can do about.

I hope this helps.
Yeng Ting

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