Introduction Throughout history, with the advance of medicine, extraordinary amount of awareness was developed about the importance of assessing erythrocyte antigens and their antibodies. Two most important systems of blood groups are ABO and Rh. The Rh system is the biggest and most complex system of blood groups. Today, from that system, five antigens- C, c, D, E, e, are by routine most frequently determined. Antigen D is a protein which is located within the membrane of the erythrocyte, and the presence of which determines whether a person is Rh-D positive or negative. If the mother is Rh-D negative, and the child Rh-D positive, hemolytic disease of the newborn may develop. It occurs because of accelerated decomposition of fetal erythrocytes caused by the mother's antibodies. For determination of hemolytic disease, the most important tests are: the test for Rh-D antigen and the direct antiglobulin test. Objective The objective of this paper is to point out the importance of precise, correct and reproducible determination of Rh factors and direct antiglobulin test for the newborn. Data sources and methods Correct record keeping is the foundation of good laboratory practice. Laboratories continuously conduct internal and external controls to keep the quality of their work on the best possible level. Samples from which the tests are derived for determining Rh-D antigens and DAT can be: the blood from the umbilical cord and the blood from test tubes with EDTA anticoagulant. Methods used are based on determining antigens on the membranes of erythrocytes. With assessing Rh-D antigen, anti-D test serum agglutinates erythrocytes if there are antigens on them. Positive reaction is established with the presence of the agglutinate, which means that the patient is Rh-D positive. With the direct Coombs test, positive reaction points to binding of antibodies or proteins of the complement system on the surface of the child’s erythrocytes. Conducting of these tests is of exceptional importance for the prevention of occurrence of hemolytic disease of the newborn, which can develop as a causal-consequential link between the Rh-D negative mother and the Rh-D positive child. In that case, the previously immunized expectant mother creates antibodies as a reaction to the child’s erythrocyte antigens which pass through the placenta and destroy fetal erythrocytes. A great amount of hemoglobin is released, which cannot be excreted, inducing anemia and hydrops of the fetus. The clinical picture can differ and prevention is of great importance by means of getting prophylaxis within 72 hours after birth. Discussion In the discussion, the case of an expectant mother who is received at the department of transfusion medicine is presented. Conclusion Precise, correct and reproducible assessments of Rh factor and direct antiglobulin tests are of extreme importance to newborns. Tests are mandatory, painless and free of charge so the obligation of every expectant mother is to go regularly for controls and tests, to assure herself and her child of the best possible healthcare.