A potential thrombus diagnosis reagent based on P-selectin monoclonal antibody SZ-51 light chain.
BACKGROUND
P-selectin is a well characterized platelet adhesion molecule that can shift from the secretory granules to the surface of activated platelets, which makes it a potential target in thrombus diagnosis and therapy. SZ-51 is a monoclonal antibody against P-selectin.


METHODS
To build a potential thrombus diagnosis reagent, we expressed the light chain of SZ-51 (SZ-LC) in P. pastoris and the protein was highly purified by the procedure combining nickel affinity purification, Q-column and Superdex 75 column chromatography. The purified recombinant SZ-LC was labeled with (99m)Tc and used for blood clearance, in vitro platelet binding and dog thrombus binding assay.


CONCLUSIONS
The yield of SZ-LC by the expression and purification method reached above 70 mg/L culture. We found that the nucleotide from (99m)Tc-SZ-LC was removed quickly through animal kidney, and (99m)Tc-SZ-LC could bind specifically to the activated human platelet in vitro. More importantly, with this recombinant protein, we successfully detected the fresh thrombus that was induced in dog vein. These results suggested that the recombinant SZ-LC expressed by P. pastoris was functional active and a potential reagent for thrombus diagnosis.
Xu
Yuanping
Y
National Key Laboratory of Protein Engineering, College of Life Sciences, Peking University, 100871, People's Republic of China.
Li
Jie
J
Fang
Wei
W
Yu
Meimin
M
Ru
Binggen
B
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology*; D055592:Biophysical Phenomena; D001792:Blood Platelets / Q000378:metabolism*; D004285:Dogs; D005822:Genetic Vectors; D006801:Humans; D007202:Indicators and Reagents / Q000378:metabolism*; D008297:Male; D019007:P-Selectin / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D010843:Pichia / Q000235:genetics; D011817:Rabbits; D019275:Radiopharmaceuticals; D011994:Recombinant Proteins / Q000276:immunology; D017263:Technetium Tc 99m Mertiatide; D013927:Thrombosis / Q000175:diagnosis* / Q000000981:diagnostic imaging / Q000235:genetics / Q000276:immunology; D015899:Tomography, Emission-Computed, Single-Photon
D000911:Antibodies, Monoclonal; D007202:Indicators and Reagents; D019007:P-Selectin; D019275:Radiopharmaceuticals; D011994:Recombinant Proteins; D017263:Technetium Tc 99m Mertiatide