Background: Study of hormonal imbalance and its implications in female infertility are an interesting area that requires to be explored in recent time. Hormonal imbalance can associated with irregular menstrual cycle, Amenorrhea, obesity and infertility in women. Other medical conditions such as polycystic ovarian syndrome, Endometriosis, stress, sexually transmitted diseases and chromosomal anomalies may be responsible for infertility in females. Objective: The aim of the present study was to evaluate the serum levels of Follicle Stimulating hormone (FSH), Luteinizing hormone (LH) and Prolactin hormone in infertile women that were referred from different infertility clinics and centres. Materials and Methods: This study comprises total 176 female subjects with age ranging from 20 to 40 years and divided in two groups. The total number of 88 infertile women along with 88 fertile women as controls was included for the present study. Serum FSH, LH and Prolactin levels were estimated by enzyme-linked immunosorbent assay (ELISA) methods. Results: The results showed maximum infertile women were found between the age group of 30-40 years. The Serum FSH, LH and Prolactin levels among infertile women was 8.77±4.65, 7.64±5.16 and 18.59±7.50 respectively. Whereas, levels of FSH, LH and Prolactin in fertile women showed that 6.71±4.12, 5.66±3.17 and 13.44±5.82 respectively. Conclusion: In this study, we found that the hormone levels have statistically significant with female infertility. The elevated levels of FSH, LH and Prolactin may be one of the important causes for infertility in women.