Probiotics, defined as live microorganisms conferring health benefits, have gained global popularity for their role in promoting host well-being. Lactic acid bacteria, known inhabitants of the gastrointestinal tract, are investigated for their probiotic properties, including acid and bile salt tolerance. Numerous researchers have documented the potential probiotic properties of lactic acid bacteria, attributing them to their ability to enhance the host animal's immune system and promote microbial equilibrium in the gastrointestinal tract. This study aimed to assess the probiotic potential of lactic acid bacteria obtained from the intestinal tract of the freshwater fish, Labeo rohita (Rohu). The isolated lactic acid bacteria underwent thorough characterization and identification, employing morphological, cultural, and biochemical analyses. Subsequently, their probiotic potential was evaluated through acid tolerance and bile salt tolerance studies. This investigation revealed that two Lactobacillus species isolated from the gut of Labeo rohita met the essential criteria for probiotics, demonstrating resilience to high acidity within the pH range of 3 to 5 and tolerating bile salt concentrations up to 0.3%.