Cyclic adenosine monophosphate (cAMP) is a ubiquitous second messenger produced from adenosine triphosphate (ATP) and involved in many cellular processes. In humans, cAMP is produced from two types of adenylyl cyclases: G protein-regulated transmembrane adenylyl cyclases (tmAC; ADCY1-9) and bicarbonate- (HCO₃^-) and calcium- (Ca²⁺) regulated soluble adenylyl cyclase (sAC; ADCY10). sAC is molecularly and biochemically distinct from other mammalian nucleotidyl cyclases. In most mammals, sAC arises from a single gene which is predicted to generate multiple isoforms via alternative splicing. In rodents, there are two molecularly identified splice variants: the "full-length" isoform (sAC_fl) and a "truncated" isoform (sAC_t). To date, biochemical and structural characterization of sAC has focused almost exclusively on the sAC_t isoform. Longer sAC isoforms, including the longest known sAC_fl, contain additional presumptive regulatory domains which have not yet been functionally characterized. Thus far, studies have been limited by the inability to obtain sufficient sAC_fl protein to allow in vitro biochemical characterization. Here, we describe attempts to heterologously express and purify human sAC_fl as well as generation of a novel genetically modified mouse strain which permits biochemical separation and purification of endogenously expressed mouse sAC_fl and sAC_t. We use these heterologously expressed and endogenous proteins to compare and contrast the biochemically properties of human and mouse sAC_fl and sAC_t.