Recombinantly Expressed Tagged SUrface Protein (RETSUP) assay: a new diagnostic system for the detection of antibodies to platelets.
J Thromb Haemost
; 22(4): 1187-1201, 2024 Apr.
Article
in En
| MEDLINE
| ID: mdl-38184205
ABSTRACT
BACKGROUND:
Current assays for the detection of (allo)antibodies to platelet antigens are often laborious and widely based on the presence of well-characterized donor platelets.OBJECTIVES:
To develop an easy-to-perform, sensitive, and specific test for the detection of antibodies against platelet antigens, in particular, glycoprotein (GP) antigens, called "Recombinantly Expressed Tagged SUrface Protein" (RETSUP) assay, which does not require donor platelets.METHODS:
Twin-Strep-tagged GP complexes were recombinantly expressed in human embryonic kidney 293 cells after stable transfection. These cell lines were used as antigen sources in the RETSUP assay, combining cell-based and enzyme-linked immunosorbent assay-based assay procedures. The assay performance was tested with recombinant antibodies, anti-human platelet antigen (HPA) reference plasmas, and anti-HPA patient sera.RESULTS:
Human embryonic kidney 293 cell lines stably expressing either Twin-Strep-labeled GPIa/IIa, GPIIb/IIIa, GPIb/IX, or GPIb/IX/V complexes or GPV as well as the distinct HPA-1, HPA-3, and HPA-5 epitopes were successfully generated. Applying the generated GP-expressing cell lines, the developed RETSUP assay proved very sensitive and specific with recombinant antibodies targeting different GPs and human plasma/serum samples. The results of the test were not affected by the GP carrying the Twin-Strep-tag or by using freshly harvested or cryopreserved cells.CONCLUSION:
The RETSUP assay is an easy-to-perform, sensitive, and specific assay for the detection of plasma/serum antibodies to platelet GP, with performance comparable to or better than those of current state-of-the-art assays in antiplatelet antibody diagnostics. Owing to the recombinant nature of the target antigens, it can be easily adapted to detect antibodies in other antibody-mediated diseases.Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Antigens, Human Platelet
/
Membrane Proteins
Type of study:
Diagnostic_studies
Limits:
Humans
Language:
En
Journal:
J Thromb Haemost
Journal subject:
HEMATOLOGIA
Year:
2024
Document type:
Article