Objective. This study describes and analyzes the relationship between political ideology, the intolerance of ambiguity, homophobia and prejudice against transgender groups in a sample of university students and graduates from the city of Lima (N = 144). Method. A quantitative descriptive-correlational study was carried out. Spanish translated versions of RWA and SDO scales were used. Also were used an adapted version of MSTAT-II (Tolerance of Ambiguity Scale) and a version of the Modern Homophobia Scale (MHS) validated for university students in the city of Lima. Results. A direct relationship was found between RWA, SDO, intolerance of ambiguity, homophobia and prejudice against certain transgender groups (transvestites and transsexuals). A multiple linear regression model suggests that RWA and intolerance of ambiguity, combined, could be applicable predictors of homophobic attitudes. Conclusion. Based on the results, it is possible to verify that there is a relationship between psychological variables that express a conservative political ideology and prejudice towards homosexuals and transgender groups, to the extent they could be considered by the most conservative individuals as very ambiguous social stimuli.