The researches find that the thrombocytopenia and bleeding are events that often occur together in neonates, this may not signify cause and effect. In fact, one study reported that 91% of neonates with low platelet counts did not develop major hemorrhage. Despite this, platelet counts are regarded as a reliable marker to determine the need for platelet transfusion. In this case study, scientists used flow cytometry as a means of determining platelet function in neonates.Most of the current methods used to determine platelet function require large volumes of blood; which is difficult to collect from premature infants.
The need for platelet transfusion
Flow cytometry can be a potent method to counter this disadvantage as it requires smaller volumes of blood; such as <500 μL. In this case study; neonates from three age groups were chosen: <29 weeks, 29–36 weeks, and >36 weeks old, and 5000 platelets were measured using FACS. One of the assays used measured how a blood sample responds to fibrinogen. Fibrinogen is a protein that promotes blood clotting by activating and binding to platelets.
To determine fibrinogen levels; 5uL of blood is incubate with various concentrations of adenosine diphosphate and its affinity for fibrinogen is test using a fluorescently label anti-fibrinogen antibody. It is find that the baseline binding to fibrinogen was below 10% for all neonates; however; the percentage of platelets that bind to fibrinogen increase when increasing levels of ADP or C-reaction proteins (proteins that are produce during inflammation or infection) are add.
Labeled anti-fibrinogen antibody
The study found a high incidence of thrombocytopenia among neonates; especially when accompanied by co-morbidities, such as sepsis. As P-selectin is a protein whose levels reflect activation of platelets; its levels were measure in this study. It is find that premature babies had less P-selectin on their surface; compare to babies born between 30–36 weeks or term babies. Also, neonates with a suspected case of sepsis or infection showed reduced activation of platelets. But This suggests that sepsis can be a potential risk factor for hemorrhage in newborns.
In many cases, there is often a delay between collecting the sample; conducting the assay, and finally analyzing it in the flow cytometer in a busy clinic. To determine if these lag periods could alter the results; samples from three healthy; individuals are take and artificial time delays are set between ;the various stages of the experiment. These included: venepuncture, stimulating the platelets, and performing flow cytometry. The baseline activation and platelet response are not alter, showing that time delays do not cause any changes to the results.