The invention provides a method for preparing stem cell-like lung cancer cells. The method comprises a step of authenticating whether the stem cell-like lung cancer cells exist in a lung cancer cell specimen to be detected or not, namely, detecting whether at least two of the following differential proteins are specifically expressed in the lung cancer cell specimen to be detected, wherein the differential proteins include protein kinase Ciota, five-time transmembrane single-stranded glycoprotein Prominin1, acetaldehyde dehydrogenase 1 or a breast cancer medicine resistance-protein ATP (Adenosine Triphosphate)-binding cassette transporter G superfamily member 2. The invention further provides a method for preparing the antigen dendritic cells loaded by the stem cell-like lung cancer cells, and a dendritic cell vaccine obtained via the method. According to the methods provided by the invention, the stem cell-like lung cancer cells can be efficiently and accurately obtained only by detecting at least two of the proteins; and the stem cell-like lung cancer cells obtained on the basis of the methods are high in purity and easy to enrich.