Introduction: The use of anesthetic gases might lead to Chromosomal Aberrations (CAs) among the operating room personnel. Aim of work: To monitor isoflurane air level in the environment of operating rooms denoting the use of anesthetic gases, and to assess the CAs among operating room (OR) personnel. Materials and Methods: Operating room personnel 184 (68 males and 116 females) were included in the study and compared with 192 other hospital areas personnel (63 males and 129 females). All subjects filled a questionnaire including personnel, family, past and present histories. All subjects were tested for chromosomal aberrations. Operating theaters were evaluated regarding the type of breathing circuits used whether open or closed, the type of anesthetic gases used, presence of air conditioning, and scavenging system. Isoflurane air level was measured using the organic method 103 by OSHA. Results: OR personnel had significantly higher percentage of CAs than control group (p value <0.001). Also it is higher with higher concentrations of waste anesthetic gases. There was a positive correlation between prolonged exposure to high concentrations of anesthetic gases and the occurrence of CAs among exposed OR personnel. Smoking had positive significant effect on frequency of CAs (p value = 0.017). Isoflurane air level was higher than NIOSH Recommended Exposure Limit (RELs) in most of measured points, and that the level is higher in dual open and closed circuits using theaters, also the level was higher in points near the anesthesia machine and in recovery rooms than critical care rooms. Conclusion: Isoflurane air level was higher than RELs in most of measured points. Personnel working in operating theaters are more at risk to develop chromosomal aberrations than controls. Recommendations: Implementation of adequate and working scavenging system and air conditioning, use of closed circuits whenever possible, periodic checking of anesthetic air levels, chromosomal studies for working personnel, and replacement of any malfunctioning parts of anesthesia machine.