Abstract Objectives To determine the influence of pH on recovery of analytes in body fluids (BFs), investigate the mechanism of pH interference, measure the frequency of abnormal-pH BFs received, and compare pH measured by meter and paper. Methods We performed pH titration in residual BFs. A low-pH BF was spiked and neutralized to investigate pH interference. We measured analytes on a Roche cobas c501 analyzer (Roche Diagnostics) and calculated the percent recovery. Measurement of pH using a meter and paper was conducted on 122 BF samples received in the laboratory. Results Enzyme activity in BFs was unaffected when pH = 7.4-8.5 lactate dehydrogenase, pH = 7.3-10.2 amylase, pH = 6.0-9.9 lipase, and pH = 1.3-11.7 all other analytes. BFs had mean (range) pH of 8.0 (5.1-8.9), with a mean (range) difference (paper ‒ meter) of ‒0.4 (‒0.6 to 1.1). Conclusions Irreversible loss of enzyme activity occurs in BFs at low pH. Few clinical BFs have pH < 7.0, but laboratories should incorporate pH measurement in BF workflows.