Background: Type 1 diabetes or insulin-dependent diabetes is rising at 2–3% per year and contributes to almost 90% of juvenile diabetes and 10% of adult-onset diabetes. The presence of autoantibodies to glutamic acid decarboxylase-65 (GAD65), insulin antibodies (IA-2), islet cell antibodies, insulin antibodies, and zinc transporter 8 (ZnT8) indicates autoimmune destruction of beta cells and thus has the highest predictive value for type 1 diabetes. The risk of developing diabetes is also higher when an individual exhibits more than one antibody. Our study aimed to compare the predictive value of diabetes positivity for a specific type of autoantibody. Materials and methods: A retrospective study was conducted at the Global Reference Lab in Mumbai over a period of 3 years (from January 2020 to July 2023) on patients and children undergoing testing for diabetes type 1 profile. Data were analyzed based on age, gender, and antibody positivity. Results: Out of the 547 patients tested, 41.68% were female, and 58.32% were male. The positivity rate for the type 1 diabetes profile was 53.75%. Glutamic acid decarboxylase-65 antibodies were detected in 45.16% of patients, followed by IA2 insulin in 15.17% of patients. 48.68% of females were positive for GAD-65 compared to 42.63% positivity in males. The prevalence of GAD-65 positivity was higher in children up to 12 years of age, at 58.86%, followed by 43.50% positivity among the 19–30 years age group. Conclusion: Our study found that more than half of the individuals (53.75%) who underwent testing, exhibited type 1 diabetes antibodies. GAD-65 positivity rates were higher in females compared to males. Clinical significance: The study provides valuable insights into the prevalence and significance of type 1 diabetes antibodies in the Indian population, offering a foundation for targeted strategies in diabetes prevention and management.