Introduction: Haemophilic animal models are used to study blood-induced cartilage damage, but quantitative and sensitive outcome measures are needed. Aim: To develop a novel quantitative method for detecting early cartilage degeneration in a haemophilic rat model of blood-induced joint damage. Methods: The ³⁵Sulphate incorporation (³⁵SO₄ ²⁻ assay) was applied to tibial and patellar cartilage of wild-type rats to quantify baseline proteoglycan synthesis and to evaluate the effect of 4-day blood exposure in vitro. Next, haemarthrosis was induced in 39 FVIII-deficient rats and characterized by changes in knee joint diameter and development of bone pathology (using micro-CT). Four- and 16-day posthaemarthrosis proteoglycan synthesis rate (PSR) was assessed using the ³⁵SO₄ ²⁻ assay, with the contralateral knee as control. Results: In vitro, a decrease in PSR in tibial and patellar cartilage was demonstrated following blood exposure. In vivo, joint diameter and development of bone pathology confirmed successful induction of haemarthrosis. In the blood-exposed knee, tibial and patellar PSR was inhibited 4 and 16 days after induced haemarthrosis. Interestingly, at day 16 the proteoglycan synthesis in the contralateral knee was also inhibited to an extent correlating with that of the blood-exposed knee. Conclusion: For the first time, early changes in cartilage matrix synthesis upon blood exposure were quantified with the ³⁵SO₄ ²⁻ assay in a haemophilic rat model, establishing this assay as a novel method to study blood-induced cartilage damage.