Flow Cytometry For Basophil Activation In Anaphylactic Shock Model Of Rats

Objective:To investigate the expression of the basophil CD63in rats’ blood with anaphylactic shock by Flow cytometry (FCM); to observe the activation of basophils and discuss the value of FCM and basophil activation test (BAT) in anaphylactic shock diagnosis.Methods:The Wistar rats of40were used, which were healthy, female, and about200g of the weight. They were randomly divided into control group and experimental group. Experimental group were classified into sensitized with no stimulate group, anaphylactic shock group,12h group after the death of anaphylactic shock,24h group after the death of allergic shock (n=8). Rats in experimental group were made into anaphylactic shock modes with OVA. The rats’ right ventricular blood and the lung tissue were extracted at different time points after the death of shock. To detect the basophil activation of rats’ blood in each group with three-color immunofluorescence marked CD63/CD45/CD203c antibody combination. The expression rates of CD63in basophil surface were gotten by CD45and CD203c shot, CD63as a basophil activation marker, which could represent the activation of the rats’blood basophils in each group. The extracted lung tissue was fixed with10%neutral buffered formalin, then was made into paraffin-embedded section and stained by HE. The lung tissue was stained by using immunofluorescence method to observe the expression of CD63in each group. The images were analyzed by BI-2000and the datas by the statistical software SPSS17.0.Results:(1) The expression rates of CD63in anaphylactic shock group,12h group after the death of anaphylactic shock and24h group after the death of allergic shock individually were17.341±2.044%,13.631±2.199%,12.236±1.646. In contrast, the control group was1.423±0.383%, and sensitized with no stimulate group was1.465±0.337%. The expression level of CD63in the anaphylactic shock group,12h after the death of shock group,24h after the death of shock respectively was significantly higher than the control group (P=0.000, P<0.01)and the difference was statistically significant. However, compared with the control group, the level in the sensitized with no stimulate group was not obviously changed. In the experimental group, the CD63expression of anaphylactic shock group come out first. With the extension of postmortem time, the CD63expression rate gradually decreased.(2) Under the microscope, CD63-positive staining was observed green and could be observed in all the anaphylactic shock death group, which indicated that the basophil cell activated. With the extension of time, positive staining gradually reduced.Conclusion:(1) In anaphylactic shock model of rats, there is abnormal basophil activation.(2) CD63, as basophil specific activation marker in Ⅰ type allergic reactions, can be a detection index for the diagnosis of allergic diseases.(3) The basophil activation test is expected to serve as a supplement in allergic reactions, and provide new method for clinical diagnosis and forensic identification of anaphylactic shock.