Increasing throughput of manual microscopy of cell suspensions using solid medium pads

Abstract

Microscopy of multiple samples using slides and coverslips is time consuming and good images are sometimes difficult to obtain due to cell movement. Our method involves manual spotting of multiple cell samples onto solid-medium pads, which creates the following benefits: •Rapid, high-quality imaging of multiple samples (hundreds per day) by visible and fluorescence microscopy.•No need for expensive automated equipment, multi-well plates or large amounts of consumables.•Wide range of working cell concentrations. The method was implemented for S. cerevisiae yeast, however it is likely to be applicable to all types of microorganisms and possibly other microscopic samples such as pollen. The lack of need for automated equipment may also make this method useful for field work.

Keywords: Agar; High throughput manual microscopy using solid-medium pads; High-throughput; Immobilization; Microbe; Microscopy; Solid medium; Yeast.

Graphical abstract

Fig. 1

Preparation of the solid medium pad using the casting cassete. Schematic of the gel preparation procedure. Numbering of the steps is identical to the description in the main text.

Fig. 2

Representative images obtained using the solid agar pad. Cells of the BY4741 Ssa1-GFP strain [6] (Mata his3Δ1 leu2Δ0 met15Δ0 ura3Δ0 Ssa1-GFP::HIS3(S. pombe his5)) were spotted onto 3% agar at different densities (A, B) or 2% agar containing 1 M potassium chloride (C), which causes formation of small foci (article in preparation). Imaging was performed using an inverted Invitrogen FLoid (20x NA 0.45 objective lens) for (A) and (B), and an upright Zeiss AxioSkop 40 with a 100x NA 1.2 oil immersion objective lens for (C).