Thiazole orange: a complement to established methods for diagnosing inherited dense granule defects.

Abstract

Inherited delta storage pool deficiencies are likely underdiagnosed due to limitations in available diagnostic tools. Achieving accurate diagnosis is thus challenging for clinical laboratories highlighting the need for improved methods. Our aim was to explore thiazole orange (TO) as a flow cytometric marker for dense granules and compare it with mepacrine, reported to have low positive predictive value, after optimization of both protocols. TO, typically used for reticulocyte marking, binds dense granules at high concentrations producing a strong signal in resting platelets. Upon activation, the signal substantially decreased due to degranulation, which was less pronounced for mepacrine. Gradual dense granule release could only be followed with TO indicating that solely TO can distinguish quantitative defects from release defects. In a small pilot study TO staining was compared to light aggregometry and flow cytometric CD63 exposure in patients with suspected inherited platelet disorders. Of 15 samples, five showed abnormal TO results; in three, the other methods agreed with granule defects, but solely TO differentiates quantitative defects from release defects. Of note, two samples showed slightly decreased TO staining, despite normal aggregometry and CD63 exposure. TO is a promising diagnostic tool for dense granule defects and may be a valuable complement in the diagnostic workup for inherited platelet disorders.