Effective estimation of correct platelet counts in pseudothrombocytopenia using an alternative anticoagulant based on magnesium salt

Abstract

Pseudothrombocytopenia remains a challenge in the haematological laboratory. The pre-analytical problem that platelets tend to easily aggregate in vitro, giving rise to lower platelet counts, has been known since ethylenediamine-tetra acetic acid EDTA and automated platelet counting procedures were introduced in the haematological laboratory. Different approaches to avoid the time and temperature dependent in vitro aggregation of platelets in the presence of EDTA were tested, but none of them proved optimal for routine purposes. Patients with unexpectedly low platelet counts or flagged for suspected aggregates, were selected and smears were examined for platelet aggregates. In these cases patients were asked to consent to the drawing of an additional sample of blood anti-coagulated with a magnesium additive. Magnesium was used in the beginning of the last century as anticoagulant for microscopic platelet counts. Using this approach, we documented 44 patients with pseudothrombocytopenia. In all cases, platelet counts were markedly higher in samples anti-coagulated with the magnesium containing anticoagulant when compared to EDTA-anticoagulated blood samples. We conclude that in patients with known or suspected pseudothrombocytopenia the magnesium-anticoagulant blood samples may be recommended for platelet counting.

Keywords: EDTA; anticoagulation; citrate; magnesium; platelet aggregation; pseudothrombocytopenia.

Fig 1

Platelet counts in 44 patients with EDTA-induced PTCP (open rhombus) corrected by parallel measurement in magnesium-anticoagulated blood samples (filled squares). The difference is statistically significant (paired t-test). In three individuals the analyser flagged the platelet count for aggregation although the values appeared to be in the normal range. This is due to the fact that PTCP is time-dependent and platelets were measured relatively early after blood sampling.

Fig 2

Time-dependent progression of anticoagulant-induced pseudothrombocytopenia (PTCP). Both cases were documented independently from the primary study. The black square and the hatched area indicate the first platelet count performed in an EDTA-anticoagulated blood sample drawn at admission and measured after arrival of the sample at the routine haematological laboratory. The red line (cross symbols) represents the time-dependent decrease of platelets in the EDTA-anticoagulated sample; the green line indicates platelet counts in citrate anticoagulated blood and the blue line (rhombus symbol) demonstrates platelet counts in magnesium-anticoagulated blood. EDTA, ethylenediamine-tetra acetic acid; TE, Thromboexact™.

Fig 3

Platelet aggregation in a blood smear from a patient with EDTA-induced pseudothrombocytopenia (left) as compared to a smear prepared from magnesium-anticoagulated blood from the same patient (right). EDTA, ethylenediamine-tetra acetic acid; TE, Thromboexact™.

Fig 4

Multiplate^R platelet function analysis. Platelet aggregation induced by adenosine diphosphate (ADP) (A, B) or arachidonic acid (ASP) (C, D) in whole blood samples from a healthy donor with normal platelet count (320 × 10⁹/l). Blood was drawn into collection tubes containing hirudin (A, C) or magnesium (B, D). The areas under the curve (AUC) are expressed by arbitrary units (u). The ADP-induced aggregation (hirudin tubes) gave an AUC of 81 u (53–122 u), the ASP-induced aggregation gave an AUC of 110 u (75–136 u). The corresponding results in blood samples collected into Thromboexact™ tubes were 10 u and 25 u, respectively.