Detection of mast cell degranulation by fluorescent molecular diffusion dynamic measurement

AIM：To establish a rapid in vitro method for mast cell degranulation tracing by raster image correlation spectroscopy (RICS). METHODS：RBL-2H3, a basophilic granulocyte mast cell line transfected with CD63-GFP plasmid, was used for evaluating the methods, including β-hexosaminidase (HEX) colorimetric assay, scanning electron microscopy (SEM) and RICS in the detection of mast cell degranulation. The sensibilities of these methods were compared. RESULTS：The sensitivities of β-HEX colorimetric assay and SEM were 5 mg/L and 3.9×10-2 mg/L, respectively. RICS detection showed obvious decrease in the diffusion coefficient at dose of 3.9×10-2 mg/L. CONCLUSION：Fluorescent molecular diffusion dynamic measurement can be used for rapid tracing of allergic substances in vitro. According to the results, RICS can achieve nearly the same extent of sensitivity as the SEM does and is far more sensitive than β-HEX colorimetric assay. Compared with SEM, RICS has several advantages: it is faster, simpler and cheaper; it can be used in living cells; it is more suitable for rapid in vitro allergenic compounds tracing. Therefore, RICS is applicable in clinic allergic antigen screening and may also be used in pharmaceutical quality control.