Genpure uv toc xcad plus

P. putida 6157 (RIKEN, Saitama, Japan) was used for biofilm cultivation. All chemicals used for the LB media were purchased from Becton Dickinson (Sparks, MD, USA). The MALDI matrices, 2,5-dihydroxybenzoic acid (DHB), and α-cyano-4-hydroxycinnamic acid (CHCA), were purchased from Sigma-Aldrich (St. Louis, MO, USA). Methanol was purchased from WAKO Pure Chemical Industries (Osaka, Japan). Ultrapure water was obtained using a Genpure UV-TOC xCAD Plus (Thermo Fisher Scientific, Waltham, MA, USA).

For sample treatment, third instar larvae at similar developmental stages (within 6 hours difference) were picked and dipped in 70% ethanol before embedding. Then, they were embedded in 10% gelatin on base-mold (7 mm × 7 mm × 5 mm; FALMA, Tokyo, Japan)^{23 (link),24 (link)}. Gelatin powder (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in ultrapure water (Genpure UV-TOC xCAD PLUS, Thermo Fisher Scientific, Waltham, MA, USA) and mixed for 10 min at 42 °C, using a thermomixer (Eppendorf, Hamburg, Germany) until a homogeneous solution was obtained^{24 (link)}. After aligning the larvae per their body axes in the embedding material, the molds were frozen rapidly in liquid nitrogen. The frozen blocks of gelatin with flies were placed in cryomicrotome (CM1950, Leica, Wetzlar, Germany) at −20 °C for 1 hour^{24 (link)}. These blocks were then placed on a tissue-holder, fixed with an optimum cutting temperature polymer (Leica, Wetzlar, Germany), and sliced at 30-μm thickness. The tissue sections were thaw-mounted onto Indium-tin-oxide (ITO)-coated glass slides (100 Ω/m² without anti-peeling coating, Matsunami Glass, Osaka, Japan) by warming them up by placing a finger on the posterior side of the glass slides^{24 (link)}. The glass slides were then dried at 40 °C for 5 min and placed in 50-mL conical tubes containing silica gel^{24 (link)}.