Wallach's Interpretation of Diagnostic Tests: Pathways to Arriving at a Clinical Diagnosis (1131 page)

   Platelets cannot be accurately counted after being stored at 4°C for more than 24 hours. In some cases, and for no known reason, the EDTA used for anticoagulation of the CBC may clump platelets, reducing their number. In such situations, the blood must be drawn with a different anticoagulant, usually 3.2% sodium citrate. A similar situation resulting in low counts is platelet satellitism (platelet adherence to neutrophils).

   Other sources of error, especially in automated counters, are giant platelet (may be counted as RBC), white cell fragments, very small red cells, or red cell fragments, counted by automated counters as platelets.

PLATELET FUNCTION ASSAY, IN VITRO
*

   Definition

   
Light transmittance aggregometry (LTA)
is based on platelet aggregation to ADP and other agonists and has traditionally been the most commonly used ex vivo assessment of platelet inhibition and activity. Due to the complexity of the assay and lack of standardization between institutions, LTA is a suboptimal test to monitor platelet activity in a clinical setting, and its use is increasingly limited to clinical trials.

   Point-of-care tests for platelet reactivity have become available for the
P2Y12
assay. The ADP-P2Y12 receptor plays a central role in the activation of platelets mediated through sustained activation of the GP IIb/IIIa platelet receptor. The assay measures ADP-induced platelet activation in citrated whole blood and reports results in P2Y12 reaction units (PRU). Initial versions of the assay also reported platelet reactivity to a thrombin receptor activation protein, which served as a measure of maximal platelet activation and reported a percentage of platelet inhibition. Increasingly, cardiovascular literature is defining platelet reactivity based on PRU alone and has defined a PRU of 235–240 as a threshold for increased ischemic events. Values at or above this level in patients on antiplatelet medications are referred to as “high on-treatment platelet reactivity” and may represent suboptimal dosing or intrinsic medication resistance.

   Other in vitro assays involve (1) measuring high shear-dependent platelet function across collagen/ADP-based cartridges. It requires only 0.8 mL of blood, and its results are obtained in a few minutes. It can be performed in the laboratory or as a POC test but lacks the reproducibility of other modalities. (2) Flow cytometric analysis of the vasodilator-stimulated phosphoprotein
(VASP),
an intracellular marker of the residual P2Y12 reactor activity and has correlated with ischemic risk in clinical trials.

   Use

   In vitro platelet function assays are increasingly used in cardiovascular medicine to assess adequate inhibition to reduce ischemic events, or conversely, when platelet inhibition is low enough after cessation of antiplatelet medications to allow for invasive procedures that carry substantial bleeding risk (i.e., noncardiac procedures).

   Although no absolute consensus for a definition of a high-on treatment threshold for platelet activity exists, it is generally accepted as (1) >235–240 PRU by VerifyNow P2Y12 assay, (2) PRI >50% by VASP-P analysis, (3) >46% maximal 5 μmol/l ADP-induced aggregation, and (4) >468 arbitrary aggregation units/minute in multiplate analyzer.

   Functional platelet assays are also utilized for

   von Willebrand disease types 1 (results may be inconclusive in mild type 1), 2A, 2B, 2M, and 3