Objective: to analyze standard precaution adherence and associated factors of nursing workers at a university hospital. Method: this is a study of mixed methods of convergent parallel strategy, carried out in Southern Brazil. The quantitative stage had 602 participants, using the instrument of sociodemographic and professional variables and the Instrument of Variables Related to Standard Precautions, analyzed using descriptive statistics. In the qualitative stage, a semi-structured interview was conducted with 24 workers, analyzed through content analysis. Results: the data showed an intermediate standard precaution adherence. In the Individual Factors dimension, the Prevention Effectiveness Scale showed high scores and the Risk Personality, Risk Perception and Knowledge about Occupational HIV Transmission scales, intermediate scores. In the dimension Work-related factors, both in the Obstacles to Following Standard Precautions Scale and in the Workload Scale, the scores were intermediate. In the Organizational Factors dimension, low scores were found for Climate of Safety and Training in Prevention of Exposure to the Human Immunodeficiency Virus and intermediate for Personal Protective Equipment Availability. Qualitative data showed that workers often select patients who they think are at greatest risk for occupational transmission to use standard precautions. Conclusion: standard precaution adherence does not occur fully among participants. Data integration allowed to conclude that, among the main elements that influence this phenomenon, is the lack of clarity of participants as to the purpose, indication and principles of standard precautions.