Publication Details
Abstract
Background: Bacterial contamination remains a major problem in mammalian cell culture because it can alter cell physiology, medium chemistry, and experimental reliability before obvious turbidity appears. Simple early-warning biochemical markers may help detect contamination without microscopy, gel electrophoresis, or image-based confirmation.
Aim: This study evaluated whether low-level bacterial contamination in mammalian cell-culture medium could be detected early using non-image biochemical and spectrophotometric markers.
Methods: Sterile Dulbecco’s Modified Eagle Medium was experimentally exposed to low inocula of safe laboratory strains, including non-pathogenic Escherichia coli K-12 and Bacillus subtilis. Four groups were included: sterile control medium, low-level E. coli contamination, low-level B. subtilis contamination, and mixed bacterial contamination. Medium samples were incubated at 37 °C and tested at 0, 6, 12, and 24 h. pH, glucose concentration, lactate concentration, conductivity, nitrite, optical density at 600 nm, and resazurin metabolic activity were measured.
Results: At 12 h, contaminated media showed clear biochemical shifts before strong turbidity was observed. Mixed contamination reduced pH from 7.42 ± 0.03 to 6.78 ± 0.05, decreased glucose from 4.50 ± 0.18 to 2.94 ± 0.16 g/L, increased lactate from 0.42 ± 0.04 to 1.86 ± 0.13 mmol/L, and increased conductivity from 13.8 ± 0.4 to 16.7 ± 0.5 mS/cm. Resazurin reduction increased significantly in contaminated groups, especially in mixed contamination. A combined marker index using pH, glucose, lactate, and resazurin activity detected contamination earlier than optical density alone.
Conclusion: Low-level bacterial contamination produced measurable biochemical changes in mammalian culture medium before marked turbidity. This model provides a simple, low-cost microbiology study based entirely on non-image verification.