Intestinal brush border membranes from 1-day-old and 4-week-old (day of weaning) pigs were affinity labeled with an Escherichia coli heat-stable enterotoxin (STa) by cross-linking 125I-STa to receptor proteins with disuccinimidyl suberate. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography revealed that a radioactive protein with a relative molecular weight of 137,000 to 145,000 was present in both age groups. A strongly radioactive protein with an apparent Mr of 90,000 was present in the 1-day-old animals but not in those that were 4 weeks old. The major radioactive protein present in the older pigs had an Mr of 64,000 to 67,000, but this protein was missing or very weakly radioactive in the younger pigs. There was no significant difference between the groups in receptor affinity for STa, although the receptor density in the older animals was marginally significantly greater. STa-stimulated guanylate cyclase activity in membranes from 1-day-old pigs was only one-sixth that in 4-week-old pigs, although the basal and Lubrol PX-stimulated activities were similar.