<p>The poultry industry is playing a vital role to meet the protein demand of highly populated countries at a comparatively low cost. This sector is under the threat of many infectious diseases. Vaccines are an effective way to overcome the diseases but repeated vaccination increases the cost for a farmer and also induces stress on birds. Due to lack of literature on multivalent vaccine of fowl cholera (FC) and avian influenza (AI) disease, the present study was the initiative towards preparation and evaluation of inactivated monovalent and polyvalent vaccine as immunoprophylaxis, for the first time in Pakistan to reduce the load of vaccination on birds. In this study, combine vaccine against fowl cholera FC and AI was developed and its immune potential was checked in poultry birds. Locally isolated and characterized strains of <i>P. multocida</i> and AI virus were used as antigen in the vaccine production. Inactivated oil and alum-based AI, FC and AI + FC vaccines were prepared and injected in experimental poultry birds. The serum was collected on 0, 14, 21, 48 and 60 days. The birds exhibited an immune response in a multivalent formulation (both oil and alum based) with the peak antibody titer (HI titer 256 and 128; ELISA titer 2.752 and 1.805) at 48th-day post-vaccination. The titers in a combined vaccine were significantly higher than the corresponding monovalent formulation. Thus oil-based combined vaccine provides a better immune response. It would be useful in minimizing cost and vaccination frequency, leading to less stress and higher immune response in birds.</p>