Figure 3: Comparison of cells cultured on TCPS and pNIPAm plates and subsequent isolation.

(a) Effect of harvesting technique (cells kept on TCPS and harvested using EDTA treatment plus scraping, versus cells kept on pNIPAm and harvested by cooling down the cultures) on the percentage of dead cells in the isolates determined by flow cytometry analysis of cells stained with ethidium homodimer. Data from 3 independent experiments are presented as mean ± sdev. *Significantly different from TCPS (p < 0.05). (b) Effect of harvesting technique on the number of attached cells, determined by the quantification of DNA 24 h after reseeding. Data from 4 independent experiments are presented as mean ± sdev. *Significant difference between pNIPAm and TCPS in paired comparison (p < 0.05). (c) Effects of surfaces and harvesting technique on THP-1 polarisation relative to resting state cells (M(−)) on TCPS of the same dataset taking quantity of CD197 (A) and CD206 (B) mRNA relative to GAPDH as index. Data of TNFα, CCL22, IL-10, and CXCL10 as well as full data of CD206 and CD197 are shown in Supplementary Fig. 2. Dashed lines represent the control level, i.e. 0 being the −ΔΔCt of M(−) cultures on TCPS. Data are presented as mean ± sdev over 3 independent experiments. *significantly different from identically treated cultures but cultured on TCPS surface. §significantly different from M(−) state cultured on TCPS of the same culture time and dataset. ⁺Significant difference in values comparing identically treated cultures that in the period 24 to 48 h after start of the polarization step are kept in polarisation medium (pol) instead of in basal medium (basal) ^#significantly different from not harvested, identically treated cultures on TCPS (p < 0.05).