The contribution of microbial amino acids through caecotrophy to tissue protein metabolism was investigated in lactating does. Attempts were made to vary microbial supply through a dietary antibiotic, Zn bacitracin, and to vary tissue demand through manipulation of litter size. Three groups of eight New Zealand does were fed different experimental diets from day 28 of pregnancy to day 26 of lactation. The control group received the basal diet formulated to meet requirements with grass hay, wheat, soyabean meal and barley grain. The second (no antibiotic) group and the third (bacitracin; BAC) group ingested the basal diet supplemented with ammonium sulfate (5 g/kg), initially unlabelled (day 1 to day 8) then labelled with15N (day 9 to day 30), while the BAC diet was also supplemented throughout with antibiotic (Zn bacitracin; 100 mg/kg). From just after birth each group of does was subdivided into two groups, each of four females, with the litter size either five (LS5) or nine (LS9) pups. The15N enrichment in liver, milk and caecal bacteria amino acids was determined by GC-combustion-isotope ratio MS. All amino acids in bacterial protein were enriched with the (15NH4)2SO4treatment, with lysine15N enrichment significantly greater in caecal bacteria (0·23 (se0·0063) atom % excess (ape)) than in liver (0·04 (se0·0004) ape) or milk protein (0·05 (se0·0018) ape), confirming the double origin (bacterial and dietary) of tissue lysine. The contribution of microbes to tissue lysine was 0·23 (se0·006) when milk protein was used as reference.