Matrix-assisted laser desorption ionization (MALDI) operated at atmospheric pressure has been shown to be a promising technique for mass spectrometry imaging of biological tissues at high spatial resolution. Recent studies have shown several orders of magnitude improvement in sensitivity afforded by coupling with a low-temperature plasma (LTP) for postionization. In this work we report the first results from "matrix-free" imaging using our atmospheric pressure (AP) transmission mode (TM) (MA)LDI source with LTP postionization. Direct MSI analysis of murine testis with no sample preparation after tissue sectioning enabled imaging of a range of lipid classes at pixel sizes of 25 μm. We compared results from the matrix-free methods with MALDI experiments in which the matrix was applied on top, underneath, or layered as a sandwich. The sandwich preparation was found to lead to ion yields approximately 2- or 3-fold higher than the other methods, indicating that the addition of a light absorbing matrix remains beneficial. Nonetheless, LDI methods confer a range of advantages, and the sensitivity improvements provided by postionization strategies are a promising step toward high-efficiency laser sampling under ambient conditions.