If thrombocytes are counted in a hemocytometer, it is necessary to count them under the phase-contrast microscope because all thrombocytes cannot be seen in the normal light, and only a smaller number of them can be counted.
Platelets appear in normal stained blood smear as small blue or colorless bodies with red or purple granules (Pappenheim’s method).
The smears must be made from the blood taken into test tubes with anticoagulant (EDTA).
If thrombocyte aggregates are present, there is danger of making a wrong lab report with low values of thrombocytes. The cell counters cannot register large aggregates of platelets and giant platelets (>20 fl). This effect is called pseudothrombocytopenia.
Determination of the thrombocyte counts
Tthrombocyte count can be made on a hemocytometer or an electronic cell counter (impedance or optical methods). The blood sample can be platelet-rich plasma or whole blood. Estimations of platelet concentration are best determined from EDTA-anticoagulated blood, where platelets generally do not aggregate.
Platelet-rich plasma for use with electrical impedance methods may be obtained by sedimentation or by low speed centrifugation.
Falsely low platelet count may be obtained by automated methods in the presence of platelet agglutinins, adsorption of platelets to leukocytes (platelet satellitism) or if the platelets have clumped due to faulty blood collection technique.
Platelet counts done on cell counters can be higher values than those done manually (only in monocytic anemia and severe haemolytic anemia).
Normal platelets are about 1-2 µm in diameter but show wide variation in shape, from round to elongated, cigar-shaped forms. A rough estimate of the platelet count can be made on a stained blood smear. If the platelet count is normal, approximately 8-15 platelets (individually or in small clumps) should be visible in each oil-immersion field.
Manual methods often use 1% ammonium oxalate as the diluting fluid, and phase-contrast microscopy for counting platelets. The normal range for platelet counts is about 150-440 x 103/µl. Platelet counts with abnormal volume distributions or very low or very high values obtained on electronic counters should be verified by examining a properly prepared and stained blood smear, unless the reliability of the particular automated instrument has been demonstrated for these ranges.
Thrombocyte count can be determined by different methods: in the hemocytometer, by an automated electronic counter, and by inspecting the blood smear. Hematologic counters use impedance method or optical method. Plasma rich in thrombocytes or whole blood can be used in the counting.
In the whole blood, impulses corresponding to the size of thrombocytes are measured. The thrombocytes count defined by automated measurements may be higher than the values obtained by the classical counting methods (only in the cases of monocytic anemia or serious hemolytic anemia).