Kozarnika cave is a renowned prehistoric site in the Balkans, which contributes significantly to our understanding of the human past due to its rich assemblages associated with the Lower to Upper Palaeolithic. Various dating methods have been employed to unravel the timing of human occupations in Kozarnika. Radiocarbon dating was used to unfold the time frame for the Kozarnikian lithic tradition uncovered in the Upper Palaeolithic sequence of the cave, and palaeomagnetic dating assigned the Brunhes–Matuyama reversal to the layer beneath the Lower Palaeolithic assemblages. In this study, we employed luminescence-dating methods, including a body of different signals to date sediment layers from the top to the bottom of the sequence covering the period of ca 30 to 700 ka. Our investigations revealed that the Kozarnikian tradition in layers 5a-c falls between 30 and 35 ka. Following that, we suggest that the Middle Palaeolithic period initiated between 250 and 309 ka and lasted until 40–53 ka. More importantly, we have updated the age of the Neanderthal radius discovered in the Mousterian assemblages to 201 ± 17 ka. Our dating resulted in a period of ca 300–700 ka for the Lower Palaeolithic assemblages in the cave. Although this age range fits perfectly with the palaeomagnetic boundary <780 ka established for Kozarnika, the possibility of reaching the threshold of luminescence dating cannot be ruled out. Thus, at this stage, this age range may represent the minimum age for the Lower Palaeolithic. •Multiple luminescence dating approaches to date a Lower to Upper Palaeolithic site.•Dating beyond the conventional limit of luminescence dating methods.•Complete chronology for the Palaeolithic site of Kozarnika in Bulgaria.•Violet stimulated luminescence (VSL) and infrared radiofluorescence (IR-RF).•Temporal constraining the prehistoric Human settlement in the Balkans.