The Conjugation of Fluorescein Isothiocyanate to Annexin V and its Application / 医学研究杂志
Journal of Medical Research
; (12)2006.
Article
in Zh
| WPRIM
| ID: wpr-564690
Responsible library:
WPRO
ABSTRACT
Objective To prepare a novel annexin V and conjugate it with fluorescein isothiocyanate and test their binding ability to phosphatidylserine(PS)exposed erythrocytes.Methods The annexin V fused with an metal chelating binding site was obtained from pichia pastoris culturing and methanol induction expression.The annexin V was purified from the culture supernatant crude product by ion-exchange chromatography and ammonium sulfate precipitation.The annexin V was conjugated with fluorescein isothiocyanate in boric buffer and the conjugate annexin V-FITC was purified by ion-exchange chromatography.Sheep red blood cells were treated with Glutaraldehyde to expose membrane PS.The annexin V-FITC binding to PS exposed erythrocytes was investigated by fluorescent microscopy.Results The novel annexin V was purified and concentrated from pichia pastoris culture by ion-exchange chromatography and ammonium sulfate precipitation.After conjugating with FITC,the annexin V was found to bind PS exposed erythrocytes by fluorescent microscopy.Conclusion The annexin V expressed recombinantly in pichia pastoris retains the binding ability to PS exposed erythrocytes and is applicable to apoptosis detection in vitro.
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WPRIM
Language:
Zh
Journal:
Journal of Medical Research
Year:
2006
Type:
Article