Three immunoglobulin G1 monoclonal antibodies, LuCa2, LuCa3, and LuCa4, were produced by fusing murine myeloma NS1 cells with splenocytes obtained from a BALB/c mouse immunized with SK-MES1 cells derived from human squamous cell carcinoma of the lung. These three monoclonal antibodies were shown to recognize different protein antigens on SK-MES1 cells by indirect immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. While the pattern of cell line distribution of antigens recognized by these antibodies was not tumor type specific, their reactivity with tissue and pleural effusion was much more informative than with cell lines. The presence of target antigens in vivo was analyzed by immunoperoxidase staining of frozen tissue sections and immunofluorescence staining of tumor cells in pleural effusions. LuCa2 antibody was reactive with lung squamous carcinoma and adenocarcinoma tumor tissues and pleural effusions, but only infrequently with those of small cell carcinoma. This antibody was also reactive with many tumor tissues from other organs as well as with various normal tissues, including alveoli and bronchus. LuCa3 and LuCa4 antibodies reacted with lung squamous carcinoma in tissues and pleural effusions, but not with lung adenocarcinoma nor with small cell carcinoma. These two antibodies reacted only weakly with normal squamous tissues of the esophagus, skin, and cervix uteri, but not with various other normal tissues. Moreover, LuCa3 had weak reactivity with squamous cell carcinoma tissue of tongue and esophagus, whereas LuCa4 had no reactivity with nonpulmonary tumor tissues. LuCa3 and LuCa4 antibodies should be of clinical interest, because our data suggest that these antibodies may be potentially useful for the diagnosis of the histological type of lung tumor cells in both cancer tissue and pleural effusions.