Lizards are seasonal breeders. Cyclic reproductive nature makes lizard as a useful model for the study of the reproductively active protein secretions in the epididymis. During breeding season, the epididymides of the lizard secret proteins that mixes with the spermatozoa and create a favourable environment for sperm maturation. In this spectrum, the aim of this study is to identify and characterize proteins which are present in the lumen of the epididymis of the lizard, E. carinata during the active phase of reproduction. The identification and analysis of the proteins are done through the proteomic approaches. The epididymal luminal fluid sample was taken from the reproductively active and inactive phase and these are subjected to the size exclusion chromatography. Two major peaks (peak 1 and peak 2) were obtained in the epididymal luminal fluid sample taken during the reproductively active phase. On the other hand, the sample from the reproductively inactive phase showed one peak (peak 1) whereas, peak 2 is not present during this phase. The peak 2 belong to reproductively active phase was later subjected to the proteomic analysis. Appropriate gel electrophoresis separation and purification methods are combined with LC-MS/MS in order to identify and characterize the proteins that are presented during the reproductively active phase. Further, in this work, nine proteins are identified including three enzymes and three heat shock proteins. Among the identified proteins, bioinformatics analysis predicts that majority of them are localized in the cytoplasm. In addition to this, an observation is made in the endoplasmic reticulum where it is seen that a close protein-protein interaction network of three molecular chaperones are involved in protein processing. Overall, this paper opens up a new dimension search for epididymal markers for the first time in reptiles, particularly lizards.