This digital resource is a scientific article published in the Journal of Cell Biology, titled 'Seeing is believing? A beginners' guide to practical pitfalls in image acquisition.' This article serves as a critical guide for researchers, particularly those new to microscopy, on common errors and challenges encountered during the acquisition of biological images. The article highlights that while modern imaging technology offers revolutionary capabilities, its increasing complexity can lead to errors if not properly understood. It reviews common practical pitfalls that can affect the interpretation of experimental data, such as issues related to sample preparation, equipment setup, and image acquisition parameters. It addresses how seemingly simple actions can introduce significant errors, for example, in colocalization analysis. This guide is invaluable for researchers across various biological disciplines who rely on microscopy for their studies. It helps them to recognize and avoid common sources of inaccuracy and bias in image acquisition, ensuring that their visual data are scientifically sound and interpretable. It encourages a deeper understanding of the imaging process beyond just operating the equipment. As a peer-reviewed publication in a reputable journal, the article provides authoritative advice on best practices in microscopy. It is particularly useful for training new users, fostering a more critical approach to image data generation, and ultimately improving the reproducibility and reliability of microscopy-based research.