Detection of cellular platinum using the monoclonal antibody 1C1

A monoclonal antibody, 1C1, developed using a trans-R,R-1,2-diamminocyclohexane (DACH) modified platinum analog (DACH-Pt-SO4) complexed with DNA was shown, using the enzyme-linked immunosorbent assay (ELISA), to have the ability to bind to both free drug DACH-Pt-SO4 and to the drug-DNA complex. Using competitive ELISA, 1C1 was found to recognize non-DACH-containing platinum compounds, such as cis-dichlorodiammine platinum (II) (CDDP). 1C1 exhibited strong binding to slot-blotted, DACH-Pt-SO4-treated DNA and moderate binding to the CDDP-DNA complex, but was unable to detect DACH containing methyliminodiacetato-trans-R,R-1,2-diamminocyclohexane platinum (II) (MIDP)-modified DNA. Immunocytochemical staining studies using 1C1 antibody on CDDP-treated BRO melanoma cells showed preferential staining of the cytosol compared with the nucleus. Although the extent of staining was dose dependent, a heterogeneous staining pattern was observed. Multicellular spheroids of MDA886LN squamous carcinoma cells treated with CDDP showed intense staining on the growing periphery compared with weak but homogeneous staining within the spheroid. Cell cycle-dependent uptake of CDDP in synchronized BRO cells may partly explain the observed heterogeneity of platinum distribution.