Rabbit anti fn polyclonal antibodies

Plasma FN concentrations were determined by an enzyme-linked immunosorbent assay (ELISA) using a well-defined domain-specific monoclonal antibody directed to cell-binding domain of FN (FN30-8; M010 TaKaRa Shuzo Co. Ltd., Shiga, Japan) as described earlier [25 (link)]. The monoclonal antibodies anti-FN were used as a coating agent. The amount of FN bound by the monoclonal antibody was quantified by rabbit anti-FN polyclonal antibodies (Sigma Chemical Co, St. Louis, MO, USA) and peroxidase-conjugated goat anti-rabbit immunoglobulins (Sigma Chemical Co, St. Louis, MO, USA) as the secondary antibodies.

The level of pFN was measured by an enzyme-linked immunosorbent assay (ELISA), using a well-defined domain-specific monoclonal antibody directed to the central cell-binding domain of FN (FN30-8; M010 TaKaRa Shuzo Co. Ltd., Shiga, Japan), as previously described [18 (link)]. The monoclonal anti-FN antibodies were used as a coating agent. The amount of FN bound by the monoclonal antibody was quantified using rabbit anti-FNpolyclonal antibodies (Sigma Chemical Co., St. Louis, MO, USA) and peroxidase-conjugated goat anti-rabbit immunoglobulins (Sigma Chemical Co., St. Louis, MO, USA) as the secondary antibodies. Each sample was analyzed twice and the concentration of pFN was calculated from the calibration curve; the mean value of the two readings was used for the statistical analysis. The pFN concentration was detected with a Synergy LX Multi-Mode Microplate Reader (BioTek, Winooski, VT, USA). A human pFN preparation (Sigma, St. Louis, MO, USA) was used as a standard for determining the FN-ELISA.

Plasma FN concentrations were determined by an enzyme-linked immunosorbent assay (ELISA) using a well-defined domain-specific monoclonal antibody directed to the cell-binding domain of FN (FN30-8; M010 TaKaRa Shuzo Co. Ltd., Shiga, Japan), as described earlier [26 (link)]. Briefly, the monoclonal antibody directed to the cell-binding domain of FN (FN30-8 M010, diluted 1:10,000) was used as a coating agent in the wells of a microtiter plate (Nagle Nunc International, Naperville, IL, USA) to bind FN from the samples. The amount of FN bound by the monoclonal antibody was quantified using rabbit anti-FN polyclonal antibodies (Sigma Chemical Co., St Louis, MO, USA, diluted 1:5000) and secondary antibody peroxidase conjugated goat anti-rabbit immunoglobulins (Sigma Chemical Co., St Louis, MO, USA, diluted 1:30,000). The test was assayed by a colorimetric reaction using o-phenylenediamine dihydrochloride/H₂O₂ as the enzyme substrate. The results were expressed in absorbance units (AU). The samples were analyzed in two different dilutions, each in duplicate. The pFN concentration is given in milligrams per liter. A human plasma FN preparation (Sigma Chemical Co., St. Louis, MO, USA, from 1.5 to 50 ng/well) was used as a standard for determining the pFN-ELISA.