66
Cytotoxicity Assay
The protocol described in Chapter II was followed.
Rosetting Techniques
The method of Jondal et aT. (67) as described in Chapter II was
used to assess the number of peripheral blood lymphocytes capable of
rosetting with sheep red blood cells.
Immunofluoresence
The methods described in Nairn (84) were followed for indirect
immunofluoresent stains of cytocentrifuge preparations of cell suspen
sions normal rabbit serum or rabbit anti-alligator immunoglobulin and a
fluorescein labeled goat anti-rabbit IgG. Immunofluorescent methods for
membrane stains are described in Chapter IV.
Hemolytic Plaque Assay
The techniques for harvesting cultured cells and assaying for
plaque-forming cells are described in Chapter II. Fresh alligator
serum diluted 1:20 was used as a complement source.
Cellular Immunoadsorbents
The method of Chess ejt al. (24) was used for fractionating alliga
tor peripheral blood lymphocytes on cellular immunoadsorbents. Rabbit
anti-alligator immunoglobulin was precipitated with 40% ammonium sulfate,
washed three times and redissolved in 0.15 M NaCl. The immunoglobulin
enriched fraction was then dialyzed against 0.15 M NaCl 0.005 M
Na^B^O^ (pH 8.3) prior to coupling onto CnBr activated Sephadex G-200
(Pharmacia). Preimmune rabbit serum, treated in an identical manner,
was coupled to Sephadex as a control.