Background and objective Early neurological deterioration (END) following acute ischemic stroke (AIS) poses a significant clinical challenge, often leading to increased disability and mortality. This study aimed to investigate the association between specific biomarkers (lactate dehydrogenase (LDH), ferritin,erythrocyte sedimentation rate (ESR),C-reactive protein (CRP), homocysteine) and the occurrence of END in ischemic stroke patients in a tertiary care hospital. Materials and methods Across-sectional hospital-basedstudy was conducted at Dr. D. Y. Patil Medical College and Hospital, Pune, from July 2022 to April 2024. Patients aged 18 and above with confirmed ischemic stroke were included, while those with hemorrhagic stroke, intracranial tumors, mass effect with midline shift, or extensive cerebral edema were excluded. Upon admission, patients were assessed using the National Institutes of Health Stroke Scale (NIHSS) and monitored daily for seven days. Biomarkers, including LDH, ferritin, ESR, CRP, and homocysteine, were measured, and patients were categorized into those with END and without END. Statistical analysis was performed using IBM SPSS Statistics for Windows, Version 20 (Released 2011; IBM Corp., Armonk, New York). Results Out of 100 patients, 12% experienced END. The mean age of patients with END was 53.50 ± 11.20 years, compared to 53.62 ± 11.74 years in those without END (p = 0.790). Males constituted 91.7% of the END group and 73.9% of the non-END group (p = 0.284). A significant association was found between END and alcohol use (41.7% in END vs. 12.5% in non-END, p = 0.010) and tobacco use (41.7% in END vs. 11.4% in non-END, p = 0.010). The mean NIHSS scores were significantly higher in the END group on day 1 (11.58 ± 3.06 vs. 7.69 ± 3.98,p = 0.002) and on day 3 (13.92 ± 3.15 vs. 6.00 ± 3.65, p = 0.001). Biomarker analysis showed a significant difference in the mean vitamin B12 levels between END and non-END groups (93.17 ± 48.58 vs. 183.45 ± 349.33, p = 0.025). Conclusion Our study identified significant predictors of ENDin ischemic stroke patients, including alcohol and tobacco use, higher initial NIHSS scores, and elevated levels of LDH, ferritin, ESR, CRP, and homocysteine, along with low vitamin B12 levels. These findings highlight the importance of routine biochemical assessments and personalized treatment plans in managing AIS, emphasizing lifestyle changes and nutritional support to improve outcomes. Limitations include the single-center design, small sample size, and lack of long-term follow-up data. Future research should validate these findings in larger, multicenter studies and explore their long-term impact on stroke recovery.