Abstract: The study aims to find out root cause for insufficient centrifugation and sample aliquoting of blood samples in clinical chemistry laboratory. Blood samples centrifuged in the laboratory were aliquoted by technicians in to 1.5 ml eppindorf cups for placement in to automated chemistry analyser. The aliquotes were allowed to settle for 1 hour and observed for presence of red rim of settled RBC at the bottom of the cups, indicating insufficient centrifugation and sample aliquoting. Frequency of such samples were analysed for time of day, type of sample tube, amount of residual serum/plasma after aliquoting, origin of sample from hospital departments. Total 13491 sample tubes analysed in 20 days at Biochemistry laboratory of New civil hospital Surat. There were 248 samples showing red rim of RBC at the bottom of the cups. Out of 248, samples received at night constituted 78%. Serum sample have 3 times chances of having insufficient centrifugation and sample aliquoting as compared to fluoride plasma. 86% of sample had residual serum/plasma of <200 ul. Neonate sample have 7 times chances of having insufficient centrifugation and sample aliquoting as compared to adult samples. 97% of samples having insufficient centrifugation and sample aliquoting were received from wards while only 3% were received from OPD. Under filling of collection tubes, neonatal samples, samples collected in plain vacuum tubes, samples collected by phlebotomist on duty at night are root causes for insufficient centrifugation and sample aliquoting.