Your browser doesn't support javascript.
loading
A simple and high -performance immobilization technique of membrane protein from crude cell lysate sample for a membrane-based immunoassay application.
Khemthongcharoen, Numfon; Uawithya, Panapat; Yookong, Nutthapon; Chanasakulniyom, Mayuree; Jeamsaksiri, Wutthinan; Sripumkhai, Witsaroot; Pattamang, Pattaraluck; Juntasaro, Ekachai; Kamnerdsook, Ampol; Houngkamhang, Nongluck; Promptmas, Chamras.
Afiliação
  • Khemthongcharoen N; Department of Biomedical Engineering, Faculty of Engineering, Mahidol University, Nakhon Pathom, Thailand.
  • Uawithya P; NECTEC, National Science and Technology Development Agency (NSTDA), Pathumthani, Thailand.
  • Yookong N; Department of Physiology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand.
  • Chanasakulniyom M; Department of Physiology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand.
  • Jeamsaksiri W; Department of Clinical Chemistry, Faculty of Medical Technology, Mahidol University, Bangkok, Thailand.
  • Sripumkhai W; Center for Standardization and Product Validation, Faculty of Medical Technology, Mahidol University, Bangkok, Thailand.
  • Pattamang P; Thai Microelectronics Center (TMEC), NECTEC, National Science and Technology Development Agency (NSTDA), Chachoengsao, Thailand.
  • Juntasaro E; Thai Microelectronics Center (TMEC), NECTEC, National Science and Technology Development Agency (NSTDA), Chachoengsao, Thailand.
  • Kamnerdsook A; Thai Microelectronics Center (TMEC), NECTEC, National Science and Technology Development Agency (NSTDA), Chachoengsao, Thailand.
  • Houngkamhang N; Mechanical Engineering Simulation and Design Group, The Sirindhorn International Thai-German Graduate School of Engineering (TGGS), King Mongkut's University of Technology North Bangkok (KMUTNB), Bangkok, Thailand.
  • Promptmas C; Thai Microelectronics Center (TMEC), NECTEC, National Science and Technology Development Agency (NSTDA), Chachoengsao, Thailand.
J Immunoassay Immunochem ; 44(1): 76-89, 2023 Jan 02.
Article em En | MEDLINE | ID: mdl-36318041
Membrane proteins are difficult to be extracted and to be coated on the substrate of the immunoassay reaction chamber because of their hydrophobicity. Traditional method to prepare membrane protein sample requires many steps of protein extraction and purification that may lead to protein structure deformation and protein dysfunction. This work proposes a simple technique to prepare and immobilize the membrane protein suspended in an unprocessed crude cell lysate sample. Membrane fractions in crude cell lysate were incorporated with the large unilamellar vesicle (LUV) that was mainly composed of POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) before coating in the polystyrene plate by passive adsorption technique. Immunofluorescence staining and the Enzyme-Linked Immunosorbent Assay (ELISA) examination of a strictly conformation-dependent integral membrane protein, Myelin Oligodendrocyte Glycoprotein (MOG), demonstrate that LUV incorporated cell lysate sample obviously promotes MOG protein immobilization in the microplate well. With LUV incorporation, the dose-response curve of the MOG transfected cell lysate coating plate can be 2-9 times differentiated from that of the untransfected cell lysate coating plate. The LUV incorporated MOG transfected cell lysate can be efficiently coated in the microplate without carbonate/bicarbonate coating buffer assistance.
Assuntos
Palavras-chave

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Proteínas de Membrana Idioma: En Ano de publicação: 2023 Tipo de documento: Article

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Proteínas de Membrana Idioma: En Ano de publicação: 2023 Tipo de documento: Article