[Real-time detection of mast cell degranulation in anaphylactoid reaction].
OBJECTIVE
To establish a new, real time, dynamic and direct optical detection method for mast cell degranulation caused by anaphylactoid reaction.


METHODS
A CD63-GFP plasmid was constructed and introduced steadily into rat basophilic leukemia (RBL-2H3) cells. The movements of CD63-GFP, which was located on both the granule membranes and the plasma membranes of RBL cells stimulated by Compound 48/80, were studied by confocal laser scanning microscope (CLSM) and total internal reflection fluorescence microscope (TIRFM) both inside and on the surface of living RBL-2H3 cells.


RESULTS
Before antigen stimulation, most granules with CD63-GFP hardly moved in RBL cells. However, after antigen stimulation, the granules moved dramatically. They reached the plasma membranes in a few minutes and fused with them instantaneously. The velocity of the granule movement toward the plasma membranes on antigen stimulation was calculated to be 0.05 micron x s(-1).


CONCLUSIONS
Analysis of the movement of each granule provided a new insight into the elementary process of degranulation. The method is rapid, sensitive and reliable, which could be used as a new detection method for anaphylactoid reaction in vitro.
Experimental Research Center, China Academy of Chinese Medical Sciences, Beijing 100700, China. jianjianghu1982@yahoo.com.cn
Jianjiang
J
Hu
Yanming
Y
Hou
Qian
Q
Zhang
Hongtao
H
Lei
Yi
Y
Wang
Danqiao
D
Wang
D000707:Anaphylaxis / Q000175:diagnosis / Q000276:immunology* / Q000378:metabolism; D000818:Animals; D015703:Antigens, CD / Q000235:genetics; D015550:Cell Degranulation; D045744:Cell Line, Tumor; D002465:Cell Movement; D008407:Mast Cells / Q000166:cytology* / Q000276:immunology*; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D010980:Platelet Membrane Glycoproteins / Q000235:genetics; D051381:Rats; D060149:Tetraspanin 30; D013997:Time Factors
D015703:Antigens, CD; C554588:Cd63 protein, rat; D010980:Platelet Membrane Glycoproteins; D060149:Tetraspanin 30