BLOOD COLLECTION TUBES

Red-top tube: Contains no anticoagulant or preservative.
Use: Serum or clotted whole blood. Serum must be separated from cells within 45 minutes to two hours depending on the test

Lavender-top tube: Contains K2 EDTA.
Use: EDTA whole blood or plasma. Separate and send plasma in a plastic transport tube labeled “Plasma, EDTA.” Send whole blood in a lavender-top tube.

Gray-top tube: Contains sodium fluoride (a preservative) and potassium oxalate (an anticoagulant).
Use: Sodium fluoride whole blood or plasma. Send plasma in a plastic transport tube labeled “Plasma, Sodium Fluoride.” Send whole blood in a gray-top tube.

Blue-top tube (also light blue-top tube): Contains sodium citrate. Be sure to use only tubes with a 3.2% sodium citrate concentration. These are easily identified by the yellow diagonal stripes on the label.
Use: Sodium citrate plasma. Send plasma in a plastic transport tube labeled “Plasma, Sodium Citrate.” Send whole blood in a blue-top tube.

Green-top tube: Contains sodium heparin or lithium heparin.
Use: Heparinized whole blood or plasma. Send plasma in a plastic transport tube labeled “Plasma, Sodium Heparin” or “Plasma, Lithium Heparin.” Send whole blood in a green-top tube.

Yellow-top tube: Contains acid citrate dextrose (ACD) solution.
Use: ACD whole blood. Send whole blood in a yellow-top tube.

Royal blue-top tube: Contains sodium EDTA for trace metal studies. Some royal blue-top tubes do not contain EDTA.
Use: EDTA whole blood or plasma. Send whole blood in a royal blue-top tube.

ABNORMAL SPECIMEN APPEARANCE

● A dark coloured urine may be positive for bilirubin or haemoglobin.

● A urine that contains whole blood may contain S. haematobium eggs.

● A black faecal specimen may contain occult blood due to gastrointestinal bleeding.

● A dark brown serum may indicate intravascular haemolysis due to sickle cell disease, severe malaria, or an incompatible blood transfusion.

● A lipaemic (fatty) serum is associated with raised triglycerides (above 3.4 mmol/l).

● A deep yellow (icteric) serum indicates that a patient is jaundiced.

● A serum sample that is abnormally viscous (thick) or turbid may contain paraproteins.

● A serum that becomes markedly turbid after being refrigerated may contain cryoglobulins or cold agglutinins.

● A blood sample that contains a high concentration of red cells from which little serum or plasma can be obtained indicates severe dehydration or a blood disorder.

CAUSES OF ERRORS OF INACCURACY (BIASE)

The causes of these can be summarized as follows:

  • Incorrect calibration of a test method.
  • Tests being read at the incorrect wavelength or An automatic pipettor is used that has been set to measure incorrect amount
  • The wrong filter being used.
  • Consistent calculation error e.g. incorrect factor is used
  • Use of unsatisfactory reagents, standards, or controls

CAUSES OF ERRORS OF IMPRECISION (SCATTER)

  • Incorrect and variable pipetting and dispensing.
  • Inadequate mixing of sample with reagents.
  • Samples are not incubated consistently at the correct temperature (when incubation of tests is required).
  • Individual items of glassware or plastic-ware are not clean or dry before reuse.
  • Equipment malfunctioning (erratic performance).
  • Incomplete removal of interfering substances, e.g. red cells when measuring analytes in serum or plasma.

WHAT MAKES A SPECIMEN TO BE REJECTED

It is unlabelled or the identity on the form does not match that on the specimen.

It is not correct for the test being requested or it has been collected in the wrong container, e.g. containing the incorrect anticoagulant.

The specimen container is leaking.

There is evidence of contamination.

A blood sample appears haemolyzed or an anticoagulated specimen contains clots.

There is too long delay in the specimen reaching the laboratory or it has not been transported

FACTORS THAT AFFECT THE MEASUREMENT OF ENZYME ACTIVITY

Temperature

Reaction rate increases with temperature. Most enzymes show their optimal activity between 30 °C and 50 °C. Above 60 °C, enzyme denaturation occurs. The incubation temperatures in test methods
must be strictly followed.

Time

The substrate concentration falls with time as the product concentration increases. Timing is therefore important because substrate and product concentrations are changing all the time and it is one of
these that is being measured. An accurate timer must be used to measure the incubation time of an enzyme and its substrate.

pH

Any increase or decrease in pH away from the optimum will cause a decrease in enzyme activity. A marked change in pH can lead to the denaturation of an enzyme. The pH of buffers and substrates used in enzyme tests must therefore be correct.

Light rays

Ultraviolet light tends to inhibit enzyme activity while blue or red light tends to increase it. Samples for enzyme analysis must therefore be protected from direct light.

SEROLOGY TESTS PART 1

Hepatitis Tests

It is performed to viral hepatitis and the type of hepatitis present e.g A, B, C & D

Syphilis Tests

The method used is Venereal Disease Research Laboratories (VDRL) and the rapid plasma reagin (RPR) test. The test results are reported as none-reactive, weakly reactive or reactive. Weakly reactive results are considered positive for to the presence of syphilis antibodies. This is a screening test thus a more specific testing for final diagnosis Incase of positive result.

Mono Test

It is used to detect the infectious mononucleosis via identification of heterophile antibody.

Rheumatoid Factor

Rheumatoid arthritis is a chronic inflammatory disease that affects the joints of the body. Rheumatoid factor is antibody produced in this condition hence is tested for diagnosis.

See also:

Serology test part 2

LEISHMAN STAINING TECHNIQUE


1. Cover the blood film (preferably methanol pre- fixed, ) with undiluted stain but do not flood the slide. If using a dropper bottle count the number of drops required to cover the film.

The undiluted stain not only acts as a fixative but also partially stains the smear. This stage is required to obtain the best possible staining results.

2. Add twice the volume of pH 6.8 buffered water (i.e. twice the number of drops as stain). The stain should not overflow. Ensure the water is well mixed with the stain by blowing on the diluted stain or mixing the stain and water using a plastic bulb pipette. Allow to stain for 10 minutes (time may require adjusting).

Diluting the stain in buffered water brings about full staining of the blood cells. The exact staining time to use should be decided when a new batch of stain is prepared.

3. Wash off the stain with tap water (filtered if not clean). Do not tip off the stain, because this will leave a fine deposit covering the film. Wipe the back of the slide clean and stand it in a draining rack for the smear to dry. The blood film should
appear neither too pink nor too blue (check
results microscopically).

HOW TO MAKE A THIN BLOOD FILM


1. Make a blood spreader from a slide which has ground glass polished sides

2. Place a drop of blood on the end of a clean dry slide too large (if too large, use a drop from the excess blood to make the film).

3. Using a clean smooth edged spreader, draw the spreader back to touch the drop of blood and allow the blood to extend along the edge of the spreader. Holding the spreader at an angle of about 30, spread the drop of blood to make a film about 40–50 mm in length (two thirds of the slide)

4. Wipe clean the end of the spreader.

5. Immediately air dry the film by waving the slide back and forth. Protect the dried film from dust and insects.

When not using a frosted ended slide,
write the patient’s name and number on the dried blood at the top or along the side of the film using a lead pencil.

6. When completely dry and within a few minutes of making the blood film, fix it in absolute methanol

SERUM GLOBULIN TEST

It is done to identify abnormalities in the rate of protein synthesis and removal

What are the normal ranges for Serum Globulin

2.5g/dl – 3.5g/dl

What causes increase in serum Globulin

  • Brucellosis
  • Chronic infections
  • Rheumatoid arthritis
  • Dehydration
  • Hepatic carcinoma
  • Hodgkin’s disease

What causes decrease in serum Globulin

  • Agammaglobulinemia
  • Severe burns