or p selectin together with the constant region of a human immunoglobulin of subclass igg1 were used the genetic construct u201c elam rg u201d described by walz et al science 250 1132 1135 1990 was used to prepare soluble l selectin igg1 fusion protein briefly plasmid dna was transfected into cos 7 cells atcc using deae dextran by standard methods see ausubel et al current protocols in molecular biology john wiley new york 1990 seven days after transfection the culture supernatant was centrifuged to remove cells and cell fragments and adjusted to 25 mm hepes ph 7 0 0 3 mm pmsf and 0 02 sodium azide and stored at 4 u00b0 c the genetic construct u201c cd62rg u201d described by aruffo et al cell 67 35 1991 was used to prepare the soluble p selectin igg1 fusion protein the subsequent procedure corresponds to the preparation of l selectin igg1 described above at section a1 the genetic construct u201c cd4 igg1 hinge u201d described by zettlemeissl et al dna and cell biology 9 347 1990 was used to prepare the soluble cd4 igg1 fusion protein the subsequent procedure corresponds to the preparation of l selectin igg1 described above at section a1 a4 procedure for the hl60 cell adhesion assay for recombinant soluble adhesion molecules 1 96 well microliter assay plates nunc maxisorb were incubated with 100 u03bcl of a goat anti human igg antibody sigma diluted 1 100.