PENETRATION AND BINDING OF MONOCLONAL-ANTIBODY IN HUMAN OSTEOSARCOMA MULTICELL SPHEROIDS - COMPARISON OF CONFOCAL LASER-SCANNING MICROSCOPYAND AUTORADIOGRAPHY

Penetration and binding of monoclonal antibody (MAb) in multicell oste osarcoma spheroids have been studied by autoradiography and confocal l aser scanning microscopy (CLSM). Optical sectioning of the 3-dimension al spheroids was performed by CLSM. Owing to attenuation of fluorescen ce intensity, FITC-labelled MAb could not be detected at depths greate r than 60 mu m within the spheroids. The antibody uptake seen in autor adiographs and CLSM images 60 mu m within the spheroids were essential ly identical. MAb had reached all parts of the spheroids within 6 h. Q uantitative measurements of the fluorescence intensity of FITC-labelle d MAb seen in confocal images and measurements of MAb bound per cell u sing flow cytometry, showed that maximum uptake was reached after 6 h. The possibility to perform both quantitative and qualitative measurem ents makes CLSM a promising method for studying antibody uptake in thi ck tissue samples.