USE OF CELL ELISA FOR THE SCREENING OF NEUROTROPHIC ACTIVITIES ON MINOR CELL-POPULATIONS IN RETINAL MONOLAYER-CULTURES

In this study we describe a large-scale screening cell ELISA protocol which is suitable for the characterization of exogenic factor effects in mixed central nervous system (CNS) culture. The main novelty of the assay is that it permits the measurement of cellular responses in pop ulations comprising as little as 2-4% of the total cell number. For st andardization of the assay, we employed antibodies against opsin and m icrotubule-associated protein (MAP2) which label distinct retinal cell classes. Embryonic chick retinal neurons were grown in microtiter pla tes and directly processed for detection of antibody binding on the sa me plate. Binding of the antibodies was saturable and the ELISA signal was proportional to the number of immunoreactive cells comprising 2-4 % and 16% of the total cell number with opsin and MAP2 antibodies, res pectively. A minimum of 2000 opsin-positive cells could be reliably de termined. Using our cell ELISA protocol, we demonstrate a developmenta l increase of both cell markers which reflected an increase in the num ber of opsin-positive cells but an enhanced expression per cell in the case of MAP2. We also show that growth-promoting activity-the presume d chick ciliary neurotrophic factor (CNTF)-stimulated the expression o f opsin in retinal cultures (EC50: 2.3 pM) and that a corresponding ac tivity is specifically expressed in the developing retina. Our results show that the cell ELISA protocol allows the rapid screening for dist inct, low-percentage cell populations responding to exogenous factors in mixed CNS cultures. (C) 1997 Elsevier Science B.V.