7
depletion of A2B5(+) cells. Similarly, visual densities of
A2B5(+) neurons and neurofilaments were equivalent in
purified and unpurified cultures. New DNA synthesis was not
required for either modulation of surface A2B5 antigen or
differentiation of cells into neurons.
Materials and Methods
Animals
White Leghorn chick embryos were used throughout this
study. Fertilized eggs were purchased from the Division of
Poultry Science, University of Florida and stored at 15C
until initiation of incubation at 37.5C in a standard
humidified egg incubator. Time in days of incubation was
used as the index of developmental age. For the present
study, 7 and 8 day embryonic optic tecta were dissected at
the tectal commissure and isolated free of non-neural
tissues aseptically in calcium-magnesium free Tyrode's
solution (CMF; Linser and Moscona, 1979).
Cell Culture
Dissociated cells were prepared by incubating tecta
for 30 min. in 0.4% trypsin (Nutritional Biochemicals,
Cleveland, OH) in CMF at 37C, followed by dissociation
with a Pasteur pipette in Medium 199 (Hank's salts,
Degenstein formula; KC Biological, Lenexa, KS) containing
0.3 mg/ml soybean trypsin-inhibitor (Sigma Chemical Co.,
St. Louis, MO) and 0.03 mg/ml DNase I (Sigma) (SBTI-DNase).