A sample-preparation-free, point-of-care testing system for in situ detection of bovine mastitis

We present a highly integrated point-of-care testing (POCT) device capable of immediately and accurately screening bovine mastitis infection based on somatic cell counting (SCC). The system primarily consists of a homemade cell-counting chamber and a miniature fluorescent microscope. The cell-counting chamber is pre-embedded with acridine orange (AO) in advance, which is simple and practical. And then SCC is directly identified by microscopic imaging analysis to evaluate the bovine mastitis infection. Only 4 μL of raw bovine milk is required for a simple sample testing and accurate SCC. The entire assay process from sampling to result in presentation is completed quickly within 6 min, enabling instant "sample-in and answer-out." Under laboratory conditions, we mixed bovine leukocyte suspension with whole milk and achieved a detection limit as low as 2.12 × 10⁴ cells/mL on the system, which is capable of screening various types of clinical standards of bovine milk. The fitting degrees of the proposed POCT system with manual fluorescence microscopy were generally consistent (R² > 0.99). As a proof of concept, four fresh milk samples were used in the test. The average accuracy of somatic cell counts was 98.0%, which was able to successfully differentiate diseased cows from healthy ones. The POCT system is user-friendly and low-cost, making it a potential tool for on-site diagnosis of bovine mastitis in resource-limited areas.