This study investigated cell immobilization of Lactobacillus rhamnosus GG in three separate protein products: native, denatured and hydrolysed whey protein isolate (WPI). Treatments were assessed for their ability to enhance probiotic survival during storage, heat stress and ex vivo gastric incubation. Spatial distribution of probiotic cells within immobilized treatments was evaluated by atomic force and confocal scanning laser microscopy, while cell viability was enumerated by plate count and flow cytometry (FACS). Microscopic analysis of denatured treatments revealed an oasis of immobilized cells, phase-separated from the surrounding protein matrix; an environmental characteristic analogous to hydrolysed networks. Cell immobilization in hydrolysed and denatured WPI enhanced survival by 6.1 +/- 0.1 and 5.8 +/- 0.1 log10 cycles, respectively, following 14 day storage at 37 degrees C and both treatments generated thermal protection at 57 degrees C (7.3 +/- 0.1 and 6.5 +/- 0.1 log(10) cfu/ml). Furthermore, denatured WPI enhanced probiotic protection (8.9 +/- 0.2 log(10) cfu/ml) following 3 h gastric incubation at 37 degrees C. In conclusion, hydrolysed or denatured WPI were the most suitable matrices for cell immobilization, while native protein provided the weakest safeguard against thermal and acid stress, thus making it possible to envision whey protein get networks as protective substrates for cell immobilization applications. (C) 2009 Elsevier B.V. All rights reserved.