Purification and activity of human fetal hepatic stimulating substance

Table 1 Assay of human fetal hepatic stimulating substance activity by ³H-TdR incorporation method

Sample	Sample volume (μL)	Concentration of protein (mg/mL)	cpm (mean ± SD)	IS	Specific activity (IS/mg protein)	Fold of purification
Control (0.9% NaCl)	25	-	1275 ± 203	-	-	-
Homogenization	25	52.600	3162 ± 329	2.48	1.89	1.00
ETOH-ppt hHSS	25	8.280	3683 ± 401	3.03	14.64	7.74
DEAE A50 peak V	25	3.470	2920 ± 251	2.29	26.40	13.97
Nondenaturing PAGE major band	25	0.169	2039 ± 114	1.60	326.89	172.76

Any value > 1.3 is significant; P = 0.05; Nondenaturing PAGE utilized DEAE-Sephadex A50 peak V as a starting material. Its purity had been improved obviously. hHSS: Human fetal hepatic stimulating substance; PAGE: Polyacrylamide gel electrophoresis; DEAE: Diethylaminoethyl; ETOH: Ethyl alcohol; IS: Index of stimulation.

Table 2 Assay of human fetal hepatic stimulating substance by enzyme linked immunosorbent assay method

Sample	Abs at 630 nm	Abs at 280 nm	Abs at 630 nm/Abs at 280 nm	Fold of purification
Control (0.9% NaCl)	0.077	0	-	-
Homogenization	0.141	2.790	0.051	1.00
ETOH-ppt hHSS	0.159	0.964	0.165	3.24
DEAE A50 peak V	0.284	0.236	1.200	23.52
Nondenaturing PAGE major band	0.197	0.019	10.368	203.29

Abs at 280 nm is absorbent value of samples at 280 nm, which represents the concentration of the samples. Abs at 630 nm represents the hHSS content in samples. hHSS: Human fetal hepatic stimulating substance. ETOH: Ethyl alcohol; DEAE: Diethylaminoethyl; PAGE: Polyacrylamide gel electrophoresis.