AIM: To establish a method for rapid detection of HLX43 conjugated  antibody and total antibody in rat serum based on enzyme-linked immunosorbent assay (ELISA), and to explore the pharmacokinetic characteristics of single intravenous injection in rats. METHODS: Based on the detection principle of ELISA, Goat anti-Human IgG (detecting total antibody) or Anti-payload antibody (detecting conjugated antibody) was coated on a 96-well plate, and the sample to be tested was added. Goat anti-human IgG Monkey ads-HRP was used as a detection antibody for detection. After adding TMB substrate for color display for a period of time, the reaction was quenched by adding stop solution. The absorption(A) was read at double wavelength 450 nm/630 nm by using SoftMax. Reading values of A450 nm-A630 nm were used for numerical fitting, and after preliminary verification based on relevant regulatory requirements, SD rats were intravenously injected 15 mg/kg HLX43 to take blood for detection of drug concentration, and pharmacokinetic parameters were calculated. RESULTS: The linear range of the analytical method was 200-6 000 μg/mL, and the results of standard curve, precision, accuracy, hook effect, dilution linearity and selectivity (matrix effect) of the analytical method were acceptable. The pharmacokinetic parameters of HLX43 were determined by quantitative analysis of serum samples from SD rats. The t1/2 was 110.9 h for conjugated antibody and 110.6 h for total antibody , respectively, Cmax was 343.3 μg/mL and 386.3 μg/mL respectively, AUC0-inf was