eng Oriental Scientific Publishing Company Biomedical and Pharmacology Journal 0974-6242 2025-09-30 18 3 2492 2503 10.13005/bpj/3272 67895 article Mohamad Zulkefly bin Mohamad Nazri 2 Norhaida binti Che Azmi 1 Mohamad Shafiq Bin Azanan 2 Clinical and Biomedical Science, Universiti Kuala Lumpur Institute of Medical Science Technology, Taman Kajang Sentral, Kajang Selangor Malaysia. Department of Paediatrics, Faculty of Medicine, Universiti Malaya, Kuala Lumpur Malaysia. Haploidentical hematopoietic stem cell transplantation (Haplo-HSCT) serves as an effective therapeutic intervention for haematological disorders, including leukemia, lymphoma, and various benign conditions. The assessment of engraftment through the quantification of chimerism is a crucial criterion for Haplo-HSCT. This investigation was designed utilising a Human Leukocyte Antigen-A24 (HLA-A24) antibody panel for the flow cytometric evaluation of chimerism post-Haplo-HSCT. Peripheral blood mononuclear cell (PBMC) specimens were procured from both patients and their corresponding donors at the University Malaya Medical Centre (UMMC). The flow cytometric analysis quantified the proportion of HLA-A24 antigen expression across a serial dilution (0%, 25%, 50%, 75%, and 100%). The FACS Diva software (Version 6.1, BD) facilitated the data evaluation, and a standard curve graph was constructed. The data were analyzed utilising the Statistical Package for the Social Sciences Software Version 27 (SPSS Version 27), with the relationship examined through the Pearson correlation test. The findings reveal a statistically significant correlation (R² = 0.99) between serial dilution and the mean expression of HLA-A24 (p<0.05), suggesting the applicability of the standard curve. The standard curve illustrated that serial dilution exhibited a direct proportionality to the expression of HLA-A24. It is suggested that this preliminary test demonstrated sensitivity for detection of T cells, B cells, and NK cells expression, with a minimum frequency of 5%, was detectable. The HLA-A24 panel enabled the monitoring of dynamic shifts in cellular subpopulations after haplo-HSCT, thereby assisting clinicians in implementing proactive management strategies. Consequently, flow cytometric analysis is a valuable technique for assessing engraftment in haplo-HSCT. https://biomedpharmajournal.org/vol18no3/flow-cytometry-based-optimisation-of-hla-a24-chimerism-monitoring-in-haploidentical-haematopoietic-stem-cell-transplantation-a-preliminary-study/ Chimerism Flow cytometry Haploidentical hematopoietic stem cell transplantation Human leukocyte antigen-A24 Peripheral blood mononuclear cell.