The microalgae Chlamydomonas reinhardtii and Chlorella sp. CCAP 211/84 were grown autotrophically and mixotrophically and their thermoluminescence emissions were recorded above 0 °C after excitation by 1, 2 or 3 xenon flashes or by continuous far-red light. An oscillation of the B band intensity according to the number of flashes was always observed, with a maximum after 2 flashes, accompanied by a downshift of the B band temperature maximum in mixotrophic compared to autotrophic grown cells, indicative of a dark stable pH gradient. Moreover, new flash-induced bands emerged in mixotrophic Chlamydomonas grown cells, at temperatures higher than that of the B band. In contrast to the afterglow band observed in higher plants, in Chlamydomonas these bands were not inducible by far-red light, were fully suppressed by 2 μM antimycin A, and peaked at different temperatures depending on the flash number and growth stage, with higher temperature maxima in cells at a stationary compared to an exponential growth stage. These differences are discussed according to the particular properties of cyclic electron transfer pathways in C. reinhardtii.