title: Sandwich ELISA Using a Mouse/Human Chimeric CSLEX-1 Antibody.
abstract: BACKGROUND
  An assay using a mouse antisialyl Lewis X (sLeX) antibody (CSLEX-1) is used clinically for screening and monitoring patients with breast cancer in Japan. However, the IgM isoform of CSLEX-1 is not preferred for the assay because the bulkiness of IgM generally causes poor accessibility to the antigen. To solve this problem, we developed an antisLeX mouse/human chimeric IgG antibody, CH-CSLEX-1, using transgenic silkworms. The performance of a homologous sandwich ELISA of CH-CSLEX1 was then evaluated.
  
  
  METHODS
  To generate CH-CSLEX-1, we used a GAL4/UAS binary gene expression system in transgenic silkworms. The reactivities of CSLEX-1 and CH-CSLEX-1 were determined in a Biacore analysis. To confirm antigen specificity, 3 antigens [sLeX, sLeA, and Lewis Y (LeY)] were used.
  
  
  RESULTS
  CH-CSLEX-1 formed correctly as an IgG class of immunoglobulin molecule with an isoelectric point close to the predicted value. The best combination for capturing and probing in a sandwich ELISA was determined as a homologous combination of CH-CSLEX-1. The CH-CSLEX-1 assay specifically detected sLeX, but not sLeA and LeY. A correlation analysis with 107 human samples showed good concordance between the conventional CSLEX-1 assay (homologous sandwich ELISA using CSLEX-1) and the CH-CSLEX-1 assay (r = 0.98). Moreover, the CH-CSLEX-1 assay was not affected by either human antimouse IgG antibodies (HAMA IgG) or HAMA IgM.
  
  
  CONCLUSIONS
  The mouse/human chimeric antibody CH-CSLEX-1 allowed the establishment of a highly specific sandwich ELISA for sLeX that was not affected by HAMA.
authors: R&D Department, Nittobo Medical Co., Ltd., Koriyama, Japan.
authors: J
authors: J
authors: Yamashita
authors: Transgenic Silkworm Research Unit, National Institute of Agrobiological Sciences, Ibaraki, Japan.
authors: I
authors: I
authors: Kobayashi
authors: Transgenic Silkworm Research Unit, National Institute of Agrobiological Sciences, Ibaraki, Japan.
authors: K
authors: K
authors: Tatematsu
authors: Transgenic Silkworm Research Unit, National Institute of Agrobiological Sciences, Ibaraki, Japan.
authors: H
authors: H
authors: Sezutsu
authors: R&D Department, Nittobo Medical Co., Ltd., Koriyama, Japan.
authors: K
authors: K
authors: Noda
authors: R&D Department, Nittobo Medical Co., Ltd., Koriyama, Japan; ishiharah@nittobogrp.com.
authors: H
authors: H
authors: Ishihara
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis / Q000737:chemistry* / Q000276:immunology; D014408:Biomarkers, Tumor / Q000032:analysis / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000737:chemistry* / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor; D011993:Recombinant Fusion Proteins