Not all light transmission aggregation assays are created equal: qualitative differences between light transmission and 96-well plate aggregometry

Melissa V. Chan,Steve P. Watson,Timothy D. Warner,Philip D. Leadbeater

doi:10.1080/09537104.2018.1466388

Melissa V. Chan, Steve P. Watson + Show 2 more

Open Access

https://doi.org/10.1080/09537104.2018.1466388

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Abstract

In this short article, submitted as part of the review on platelet function testing, we illustrate the quantitative and qualitative differences between classical light transmission aggregometry (LTA) and 96-well plate aggregometry.We show that responses to platelet agonists and antagonists differ depending upon the method of aggregation testing. For example, in 96-well aggregometry, responses to collagen are strongly inhibited by P2Y12 receptor antagonists while in LTA they are much less affected. Furthermore, we explore the importance of differences in the mechanical environment upon platelet aggregation.We emphasize that LTA and 96-well aggregometry are not interchangeable assays. These two assays are best used as complementary tests to explore platelet function in depth.

Highlights

First reported over 50 years ago [1], light transmission aggregometry (LTA) is still considered as the gold standard way to measure platelet reactivity
Aggregation to AA was blunted by AR-C66096 in LTA while it was abolished in 96-well aggregometry
Aggregations induced by U46619 were unaffected by aspirin in either LTA or 96-well aggregometry, but were substantially reduced by AR-C66096 (Figure 1B)

Summary

Introduction

First reported over 50 years ago [1], light transmission aggregometry (LTA) is still considered as the gold standard way to measure platelet reactivity. It relies upon the simple principle that as a suspension of platelets aggregates, the transmission of light through the suspension increases allowing one to follow platelet aggregation in real time It has the advantage (which was not realized when first described) of being able to record the preceding change in shape of platelets from a discoid to a spherical structure (seen as an increase in optical density of the suspension). To perform this assay in standard light transmission aggregometers, 200–500 μl suspensions of platelets in plasma (plateletrich plasma, PRP) or in buffer (washed platelets) are placed in warmed cuvettes containing a magnetic stirrer bar and typically stirred at 1200rpm (in some aggregometers the rate can be varied) before addition of test compounds. The degree of light transmission reflects aggregation but is influenced by platelet granule secretion, and there are differences between the optical density of micro- and macro-aggregates

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