Background: The effect of direct oral anticoagulants (DOAC) on laboratory tests dependent on the production of their targets, factor IIa and factor Xa, is a well-known problem and can cause both false positive and negative results. In particular, the situation in patients who develop lupus anticoagulant (LA) antibodies is highly complex. To evaluate the effectiveness of DOAC therapy in lupus-positive patients, 31 samples were enrolled in this retrospective study. All patient samples were spiked with three types of DOAC (dabigatran, DABI; rivaroxaban, RIVA; and apixaban, API) in a concentration that significantly influenced the screening test for LA and thus can mask the presence of LA. Subsequently, the DOAC was always unbound by the DOAC-Stop procedure. DOAC levels before and after binding were determined by functional assays, followed by liquid chromatography coupled with mass spectrometry (LC-MS) analysis. Methods: The determination of DOAC levels was performed by direct thrombin assay and determination of anti-Xa activity with specific calibration as functional tests for DABI and xabans (API and RIVA). To determine concentration levels of API, DABI, and RIVA, our in-house LC-MS method was used. Results: The results of LA-positive samples show significant differences between functional tests and the LC-MS method both before and after DOAC binding. Conclusions: The acute findings of the presence of LA-type antibodies fundamentally affects the determination of DOAC by functional tests, and in this case, it is necessary to use LC-MS analysis to determine the true value. If patients treated with DOAC develop LA of medium and higher titers, we do not recommend checking DOAC levels with functional tests.