This study was conducted to find out the distribution of ABO phenotypes frequencies in urban area of Southern Punjab Multan Pakistan. A total of 1087 subjects (both male & female) were taken and their blood samples were tested for ABO blood group & Rh. by using anti sera of ABO blood grouping i.e. Anti-D, Anti-A and Anti-B, and confirmed by standard technique. Doubtful results were checked by tube method. The data was analyzed by applying Pearson correlation and chi square test. In the present study overall prevalence of blood group & Rh. positive was 997/1087 (91.72%) as compared to 90/1087 (8.28%) blood group & Rh. negative (P< 0.05). It is a part of evolution with a natural and random genetic drift selection. ABO blood group types were equally distributed but there was no significant difference in both sexes. However, among ABO blood group & Rh positive; dominance frequency was ‘B’ positive with 373 (34.32%) followed by ‘O’ positive 293 (26.95%), ‘A’ positive 218 (20.06%) and ‘AB’ positive with frequency of 113 (10.40%) respectively, and among ABO blood group & Rh. negative; ‘O’ negative was more prevalent with 42 (3.86%) followed by ‘B’ negative 23 (2.12%), ‘A’ negative 22 (2.02%), and ‘AB’ negative with frequency of 03 (0.28%) respectively In the present study ABO blood group ‘B’ positive was dominant as compared to other ABO blood groups & Rh positive (P> 0.05). Analysis of the data collected in the present study indicated that there was a dominant frequency of ABO blood group & Rh. positive as compared to Rh. negative, and blood group ‘B’ positive was dominant as compared to other ABO blood group types. The determination of the frequency of ABO blood group types in urban area Southern Punjab Multan would help us in understanding the distribution of ABO phenotypes. Whereas, the differences of ABO blood group’s frequencies in different races and regions is a part of evolution with a natural and random genetic drift selection.