title: Immunodiagnosis of platelet activation in immune thrombocytopenia through scFv antibodies cognate to activated IIb3 integrins.
abstract: Immune thrombocytopenia (ITP) is an autoimmune bleeding disorder characterized by low platelet count and presence of IgG autoantibodies to platelet surface glycoproteins, such as α IIbβ3 and GPIb/IX. Our previous work has shown that platelets in ITP patients exist in an activated state. Two different marker-based approaches are used to study the course of platelet activation: (1) binding of PAC-1 antibody, signifying a change in αIIbβ3 conformation, and (2) expression of P-selectin, signifying alpha granule content release from platelets. Here, we describe the development of a new scFv antibody (R38) that, compared with PAC-1, appears to better distinguish between platelets of ITP patients and healthy controls. Notably, R38 was generated using commercially sourced resting-state integrin that was coated on a microtiter plate. Its ability to distinguish between ITP patients and healthy controls thus suggests that inadvertent integrin activation caused by coating involves a conformational change and exposure of a cryptic epitope. This report also describes for the first time the potential use of an scFv antibody in the immunodiagnosis of platelet activation in ITP patients.
authors: a Department of Internal Medicine ; Postgraduate Institute of Medical Education and Research ; Chandigarh , India.
authors: Preeti
authors: P
authors: Bhoria
authors: b Hematology; Postgraduate Institute of Medical Education and Research ; Chandigarh , India.
authors: Neelam
authors: N
authors: Varma
authors: a Department of Internal Medicine ; Postgraduate Institute of Medical Education and Research ; Chandigarh , India.
authors: Pankaj
authors: P
authors: Malhotra
authors: a Department of Internal Medicine ; Postgraduate Institute of Medical Education and Research ; Chandigarh , India.
authors: Subhash
authors: S
authors: Varma
authors: c Immunopathology; Postgraduate Institute of Medical Education and Research ; Chandigarh , India.
authors: Manni
authors: M
authors: Luthra-Guptasarma
mesh_terms: D000328:Adult; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D018613:Microscopy, Confocal; D008875:Middle Aged; D015539:Platelet Activation / Q000276:immunology*; D010974:Platelet Aggregation / Q000276:immunology; D010976:Platelet Count; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000276:immunology* / Q000378:metabolism; D016553:Purpura, Thrombocytopenic, Idiopathic / Q000097:blood / Q000175:diagnosis / Q000276:immunology*; D012680:Sensitivity and Specificity; D057127:Single-Chain Antibodies / Q000276:immunology* / Q000378:metabolism; D055815:Young Adult
chemical_list: D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D057127:Single-Chain Antibodies
keywords: ADP, Adenosine diphosphate; ADP, scFv, ITP, PAC-1, platelets; CDR, complementarity determining region; IPTG, Isopropyl beta-D-thiogalactoside