The major use of alternative light sources (ALS) in the evaluation of bloodstains has been primarily focused on detection of whole blood, with relatively little attention to visualization of blood serum. Serum may become separated from blood pools during clotting, and because it is relatively invisible on certain backgrounds, go undetected by a perpetrator attempting to clean up a crime scene. Recently, Ultraviolet 365 (UV 365) was shown to be an effective tool in blood evaluation, useful for detection of even minute quantities of blood serum. Here the effects of environmental conditions on blood serum stain appearance were evaluated, including temperature, pH, protease sensitivity, solubility, and aging. Interestingly, it was found that the UV fluorescence of serum increases upon exposure to heat, which was accompanied by color changes under visible light and decreased solubility in multiple solvents. The efficiency of visualization of serum stains was somewhat variable, depending on the type of material on which it was dried. Finally, the current study documents the effect of heating on formation of fluorescent serum halo rings in dried bloodstains. Taken together, these data demonstrate that blood serum detection may be affected by certain conditions that influence its visualization under both visible and UV light.