Over the last 50 years the traits of valuable indigenous Lithuanian Žemaitukai horse have not been investigated and during the last decade the breed became on the verge of extinction. Recently certain measures were undertaken to preserve the breed, evaluate its present characteristics and compare them with those reported earlier. Body size and conformation traits of present-day Žemaitukai horse (ŽH) were found corresponding or similar to those of the ancient type, showing that many valuable characteristics of the breed are retained. Mares’ milk yield and composition were evaluated, as well as development of foals based on body dimensions. Genetic variation, genetic structure as well as relationship between the lines and families of ŽH were studied using blood typing and electrophoretic analysis of serum proteins. Gene frequencies are presented at six blood group (A, D, C, Q, P, K) and five protein ( Al, Es, Gc, Xk, Tf) loci. The genetic diversity within blood groups and serum proteins in ŽH kept in a closed population showed that out of eleven genetic systems examined, six were polymorphic. This is especially so for the A and D, as well as Es and Tf systems. The distribution of allele frequencies varied between the lines and families.