title: The importance of characterization of FITC-labeled antibodies used in tissue cross-reactivity studies.
abstract: Fluorescein isothiocyanate (FITC)-labeled antibodies are widely used as primary antibodies in the tissue cross-reactivity (TCR) studies for the development of therapeutic antibodies. However, the effects of FITC-labeling on the characteristics of an antibody are poorly understood. The present study was performed to examine the effect of FITC-labeling on the binding affinity and immunohistochemical staining profile of an antibody, using several antibodies with different FITC-labeling indices. The results showed that the FITC-labeling index in antibody was negatively correlated with the binding affinity for its target antigen. Immunohistochemically, an antibody with a higher labeling index had a tendency to be more sensitive, but was also more likely to yield non-specific staining. Based on these findings, we recommend that a FITC-labeled antibody used as a primary antibody in a TCR study should be carefully selected from several differently labeled antibodies to minimize the decrease in the binding affinity and achieve the appropriate sensitivity and interpretation of the immunohistochemistry.
authors.affiliation: Safety Assessment Department, Chugai Pharmaceutical Co., Ltd., 1-135 Komakado, Gotemba, Shizuoka 412-8513, Japan. takaihrt@chugai-pharm.co.jp
authors.forename: Hirotake
authors.initials: H
authors.lastname: Takai
authors.forename: Atsuhiko
authors.initials: A
authors.lastname: Kato
authors.forename: Teruo
authors.initials: T
authors.lastname: Nakamura
authors.forename: Tatsuhiko
authors.initials: T
authors.lastname: Tachibana
authors.forename: Takayuki
authors.initials: T
authors.lastname: Sakurai
authors.forename: Masahiko
authors.initials: M
authors.lastname: Nanami
authors.forename: Masami
authors.initials: M
authors.lastname: Suzuki
mesh_terms: D000345:Affinity Labels; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology; D000918:Antibody Specificity; D003429:Cross Reactions; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry; D005455:Fluorescent Antibody Technique / Q000379:methods*; D006801:Humans; D009928:Organ Specificity
chemical_list: D000345:Affinity Labels; D000911:Antibodies, Monoclonal; D016650:Fluorescein-5-isothiocyanate