Polymer microsphere flooding technology has been widely used in oil production plants in North China Oilfield and achieved good results. Due to the interference of linear polyacrylamide ( HPAM ) in oilfield produced fluid, it is difficult to accurately determine the content of polymer microspheres. In this paper, the UV spectrophotometer method was used to optimize the best detection wavelength by maximizing the slope of the linear fitting curve of different concentrations of samples at a specific wavelength. The influence of HPAM was eliminated by adding masking agent to the reference and the solution to be tested by partially masking the UV response. The quantitative detection of polymer microspheres content under HPAM interference was realized. The experimental results showed that the optimal UV detection wavelength of polymer microspheres was 196 nm. A masking agent with ammonium group was optimized. When the dosage of masking agent was 0.3 % and the microsphere concentration was 1 mg/L-200mg/L, the linear relationship between absorbance ( y ) and microsphere concentration ( x ) was y = 0.0039x + 0.0024, and the correlation coefficient was 0.9987, indicating the feasibility of the detection method. The results can provide reference for the quantitative detection of polymer microsphere content in similar reservoirs.