Investigations of Heat Shock Protein 70 for Non-Invasive Detection of Heat Stress in Dairy Cows

Abstract

Heat stress (HS) constrains dairy production, impairing welfare, milk yield, and reproduction. Common indicators—rectal temperature (RT), infrared thermography (IRT), and reticulorumen temperature (RRT)—capture environmental and physiological responses but provide limited insight into cellular stress. This thesis evaluated heat shock protein 70 (HSP70) as a non-invasive indicator of HS in dairy cows, focusing on its analytical reliability, biological variability, and temporal dynamics in pasture-based systems. The research combined a literature review with analytical validation, methodological optimisation, and seasonal and time-lag analyses. The review identified key gaps in applying HSP70 as a biomarker for HS detection. Experimental work addressed analytical challenges: an in-house competitive ELISA showed strong agreement with a commercial assay across milk, blood, and saliva, with greater precision, wider detection range, and lower cost. Sample handling studies showed that milk processing affects HSP70 stability, with skimming and short-term refrigeration or freezing improving preservation. Seasonal analyses revealed increased HSP70 concentrations in milk, blood, and saliva during elevated heat load, although milk HSP70 showed greater variability and associations with lactation stage. Time-lag analyses demonstrated that HSP70 responds more slowly than conventional indicators: RT and IRT reflected immediate heat load, RRT captured short-term internal changes, whereas milk HSP70 showed delayed responses linked to prior heat exposure. This lag aligns with the biological role of HSP70 as a stress-induced molecular chaperone requiring transcriptional activation before detection. Overall, milk HSP70 is not a real-time HS indicator but a complementary, non-invasive biomarker reflecting cumulative cellular stress. Integrating molecular and physiological indicators provides a more robust framework for HS assessment.