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Description: In vitro technique is defined as a research developed outside of a living organism and its main material for analysis is organs, tissues, and cells. This research technique is widely used in pharmacological, biological fields, and it is the precursor of clinical research, being the bottom of the evidence pyramid. Contrastingly, a systematic review (SR) is a type of research on the top of the evidence pyramid, with strict methodology and performed to answer a specific research question through the selection and assessment of primary studies. SR usually include primary studies of clinical research; however, this scenario has been changing, with SR including pre-clinical studies as in vitro ones. There is an increase in SR publications, but a decrease in methodological quality. One important step to improve SR methodology is the use of tools to critically evaluate the primary studies included. Those tools provide an outline of the quality and/or risk of bias of the research performed, and its potential in informing clinical recommendations. There is a range of tools for the assessment of clinical studies. However, there is not an appraisal tool for in vitro studies with cell culture. Usually, researchers utilize adaptations of appraisal tools developed for in vivo or clinical studies and this leads to numerous inappropriate criteria that harm the assessment of the quality of the in vitro studies. Thus, the objective of this methodological study is to identify, collect, and analyze the SR of in vitro studies with cell culture to highlight the appraisal tools utilized currently and to serve as a basis for the development of a critical appraisal tool for in vitro studies with cell culture capable of capture all the essential factors related to this type of research.