The objective of this research was to investigate the influence of pH-modification and concentration on the chymosin-induced gelation of milk. Milk was adjusted to pH 6.0 and 5.5 and then concentrated to 3× using osmotic stress. The concentrates were compared with those prepared with milk at native pH. The gelation behavior was monitored using rheology and light scattering, in situ. Partial acidification accelerated the coagulation of casein micelles and the gelation time decreased from 55 ± 2.5min to 9.0 ± 2.0 min and 6.0 ± 0.5 min at pH 6.0 and 5.5, respectively. Concentration (3×) increased the gel strength with the highest elastic value 977 ± 81 Pa was found at pH 6.0. These results confirmed previous literature data. It was demonstrated that in untreated milk, the gelation time was not affected by casein concentration, while in the case of the pre-acidified samples, the gelation times increased after concentration. This was attributed to a change in the composition of the serum phase, including an increase in soluble proteins. It was also found that more caseins were released for the pre-acidified samples during the concentration process. When the concentrated acidified casein micelles were added to untreated milk, the gels showed higher elastic modulus (around 372 ± 30 Pa) compared with control (146 ± 10Pa). This effect was not due to free calcium concentration, as when the mixtures were equilibrated against milk, no gelation was observed. The lack of gelation could not be solely attributed to changes in diffusible calcium phosphate, but to a partial solubilisation of the casein micelles. The results demonstrated that both soluble caseins and diffusible calcium phosphate play an important role in the coagulation of casein micelles induced by chymosin.