The adoptive transfer of alternatively activated macrophages (AAMs) has proven to attenuate inflammation in multiple mouse models of colitis, however the effect of cryopreservation on AAMs, the ability of previously frozen AAMs to block DNBS (Th1) and oxazolone (Th2) colitis, and their migration post-injection remains unknown. Here we have found that while cryopreservation reduced mRNA expression of canonical markers of IL-4-treated macrophages [M(IL-4)], this did not translate to reduced protein or activity and the cells retained their capacity to drive the suppression of colitis. The anti-colitic effect of M(Il-4) adoptive transfer required neither T or B cell nor peritoneal macrophages in the recipient. Following injection into the peritoneal cavity, M(IL-4s) migrated to the spleen, MLNs and colon of DNBS-treated mice. The chemokines CCL2, CCL4, and CX3CL1 were expressed in the colon during the course of DNBS-induced colitis. The expression of integrin β7 on transferred M(IL-4)s was required for their anti-colitic effect, while the presence of the chemokine receptors CCR2 and CX3CR1 were dispensable in this model. Collectively the data show that M(IL-4)s can be cryopreserved M(IL-4)s and subsequently used to suppress colitis in an integrin β7-dependent manner and we suggest that these proof-of-concept studies may lead to need cellular therapies for human inflammatory bowel disease.