The capacity for macromolecular absorption in the gastrointestinal tract of the newborn calf erodes with time after birth and ceases between 24 and 48 h of age. Little is known about mechanisms governing this process. Eight bull calves, whose births were observed, were removed from their dam before standing. Calves were slaughtered at birth or 48 h of age (n = 4/age). Calves slaughtered at 48 h received 4 L of colostrum within 4 h of birth and milk replacer at 24 and 36 h. Cross sections were collected from the duodenum midway between the pyloric sphincter and the liver. Sections were rinsed in deionized water, snap-frozen in liquid N and stored at −80°C. Samples were pooled to create 2 pools per age (0, 48) with 2 calves represented in each pool. Protein expression from 48-h pools was compared with 0-h pools using 2 2-D fluorescent gels. Proteins with approximately 2-fold greater expression from calves in one age group were further analyzed using mass spectrometry to identify unique proteins. Alcohol dehydrogenase and fructose-bisphosphate aldolase B expressions were 1.8-fold higher in calves at birth. Fifteen proteins had increased expression 48 h after birth; several identified proteins were involved in multiple physiological pathways. Functional groupings based on gene ontology were: immunity (n = 5), angiogenesis (n = 4), lipid metabolism (n = 3), protein synthesis (n = 3), nutrient delivery (n = 2), cell differentiation (n = 2), cell growth (n = 2), nervous system maturation (n = 1), and vitamin A metabolism (n = 1). Greatest increases in expression were for peroxiredoxin-6 (immunity), serum albumin (nutrient transport), and pre-mRNA processing factor 19 (protein synthesis). These proteins increased by 11.1-, 6.9- and 4.5-fold 48-h after birth, respectively. Results indicate that feeding colostrum to calves may initiate blood vessel, nervous, and other tissue development and may also be important for vitamin A metabolism. Subsequent studies aim to differentiate between effects of calf age and colostrum feeding.