Enterotoxaemia, caused by Clostridium perfringens toxins, is one of the most prevalent clostridial diseases in ruminants. The diagnosis is typically based on the detection of C. perfringens toxins in the intestinal content; however, challenges remain in achieving a practical, reliable, and definitive diagnosis. We present comparative findings of the enzyme-linked immunosorbent assay (ELISA), immunohistochemical (IHC), and pathological in 150 ruminants (sheep, goats, and cattle) suspected of enterotoxaemia. The present results revealed that 19 out of 150 ruminants were positive for at least one C. perfringens toxin (α, β, and ε) by ELISA. Moreover, our findings indicated that macroscopic and histopathological observations were congruent with clostridial enterotoxaemia. IHC for C. perfringens stained numerous long, rod-shaped bacteria present in the intestinal lesions in all toxin-positive cases (19/19). Additionally, positive immunohistochemical staining was observed in 115 of 131 toxin-negative ruminants. Our present findings suggest that elevated rates of positive immunohistochemical staining, particularly in histopathological intestinal lesions, may significantly contribute to the pathological diagnosis of enterotoxaemia. It also suggests that when toxin detection is unfeasible, enterotoxaemia can be diagnosed more effectively and securely by concurrently assessing IHC and histopathological findings, hence allowing for the identification of potential positive cases.