The ongoing increase in genomic surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) lineages remains a significant global challenge, mainly due to the incompatibility of genetic sequencing with the technological and economic capabilities of most countries worldwide. Additionally, the continued circulation and emergence of numerous lineages of the Omicron variant with some specific mutations represent a challenge in developing straightforward tests for their discrimination. In this study, we developed a multiplex assay based on reverse transcription loop-mediated isothermal amplification with lateral flow assay detection (RT-LAMP-LFA). This assay aims to enhance the accessibility of genomic surveillance and to deliver more precise epidemiological data to support public health decision-making. To demonstrate its utility, we used the assay as a proof of concept for discriminating between the SARS-CoV-2 Omicron lineages BA.1 and BA.2. When comparing the results of the new assay with the gold standard method of genetic sequencing in a panel of clinical samples, the multiplex RT-LAMP-LFA demonstrated excellent diagnostic performance. For BA.1 lineage detection, the assay achieved 100% sensitivity, 100% specificity, and 100% accuracy, while for BA.2, it showed 100% sensitivity, 95% specificity, and 96% accuracy. The overall simplicity of the method combined with advantages such as short analysis time (40 min), low cost (∼5 per test) and adaptability to the point-of-care format make the multiplex RT-LAMP-LFA assay an important screening tool for inferring SARS-CoV-2 Omicron lineage positive samples, thus alleviating the high demand for sequencing and expanding genomic surveillance, even in remote locations.