Position-referenced microscopy for live cell culture monitoring

Abstract

Position-referenced microscopy (PRM) is based on smart sample holders that integrate a position reference pattern (PRP) in their depth, allowing the determination of the lateral coordinates with respect to the sample-holder itself. Regions of interest can thus be retrieved easily after culture dish transfers from a cell incubator to the microscope stage. Images recorded at different instants in time are superimposed in a common coordinate system with subpixel accuracy. This paper presents such smart Petri culture dishes and their use for live cell culture monitoring. The impact of the PRP on the light budget is discussed and performances are demonstrated. First results on the application of PRM to the observation of apoptotic body internalization are reported.

Efferocytosis of apoptotic human papillomavirus‐positive cervical cancer cells by human primary fibroblasts

“…In addition, the monitoring of AC clearance was evaluated by position‐referenced microscopy as previously published (Galeano et al., ). By tracking the fluorescent dyes during co‐culture between PKH26‐labelled HPFs and CFDA, SE‐labelled HeLa LAC at different time periods, we observed that apoptotic corpses quickly adhered to the plasma membrane of HPFs and were efficiently internalised by HPFs several hours after first tethering (see Video S1 in the Supporting Information).…”

“…The percentages of HPFs or Mϕ having ingested AC were calculated. Uptake of AC by HPFs was also investigated by positionreferenced microscopy to track the fate of HPFs exposed to AC, as previously described (Galeano et al, 2011). Briefly, 5-(and-6)-carboxyfluorescein diacetate, succinimidyl ester-(CFDA, SE; Invitrogen) labelled AC were incubated with PKH26-labelled HPFs for 11 h with an observation every hour.…”

Visual in-plane positioning of a Labeled target with subpixel Resolution: basics and application

“…Cell culture boxes, including this new technology, have been used by biologists for researching DNA material transfers after cell death in the context of cervical cancer [19]. Also, the above described technology is used for quantifying the proliferation of human liver tumor-derived cells by means of wound healing assays.…”

Characterization and compensation of XY micropositioning robots using vision and pseudo-periodic encoded patterns

“…The reproducibility of the visual measurement has been experimentally evaluated and is better than 10 nm [19].…”