<p>Immunosenescence, a recognized hallmark of aging, is characterized by imbalances in immunocyte populations and a state of chronic inflammation. However, the tissue-specific dynamics of these changes and their potential as predictive biomarkers for aging remain poorly characterized. In this study, we established a multi-tissue immunological signature as a robust predictor of biological age by integrating immunocyte and cytokine profiling. Using Sprague–Dawley (SD) rats from five age groups (1–12&#xa0;months), we systematically quantified 45 immunocyte subsets across peripheral blood, mesenteric lymph nodes, thymus, and spleen using flow cytometry, and profiled 22 serum cytokines/chemokines via Flexible Multi-Analyte Profiling (xMAP). Firstly, classic age-dependent shifts were observed across our rat samples, including progressive thymic involution and depletion of peripheral T-cells. Cytokine levels exhibited age-related chronic inflammation progression, marked by elevated IL-1α, granulocyte colony-stimulating factor (G-CSF), and TNF-α. To integrate these multidimensional datasets into a predictive aging metric, we employed Least Absolute Shrinkage and Selection Operator (LASSO) regression, selecting 22 biomarkers through regularization (λ = 0.111). The integrated model combining cellular and cytokine data demonstrated superior performance (training R² = 0.957, validation R² = 0.887), outperforming single-modality models based on immunocytes or cytokines. Notably, splenic parameters dominated the aging signature, contributing seven biomarkers representing 60% of model weight—particularly Th-cell expansion and Tc-cell depletion. Peripheral blood Th-cell proportion emerged as another key predictor. Our findings position the spleen as a critical aging hub and identify peripheral/splenic Th-cell modulation as promising therapeutic targets for age-related immune dysfunction, revealing novel mechanistic insights into aging-associated immune remodeling.</p>