The commercially available TaqMan Gene Expression Assays Hs01650998_m1, Hs00171042_m1, Hs00171065_m1, Hs00234140_m1, Hs01567026_m1 and Hs00154355_m1 (Applied Biosystems, Foster City, CA, USA) in combination with TaqMan EZ technology (Applied Biosystems) were used to determine the expression levels of CXCL17, CXCL10, CXCL9, CCL2, CD86 and CD68, respectively. The RT–PCR profile was 50 °C for 2 min, 60 °C for 30 min and 95 °C for 5 min followed by 45 cycles of 95 °C for 20 s and 60 °C for 1 min. Emission from the released reporter dye was measured by the ABI Prism 7700 Sequence Detection System (Applied Biosystems). All qRT–PCR analyses were carried out in triplicates. The concentration of 18S rRNA was determined in each sample by real-time qRT–PCR (Applied Biosystems) for normalisation of chemokine mRNA levels (Bas et al, 2004 (link)). Chemokine expression levels in primary tumours and cell lines are expressed as relative quantity (RQ) calculated according to the equation: 2^-(Δct of the sample−the median Δct value of the normal colon tissue samples). Δct is the ct value of the chemokine mRNA minus the ct value of 18S rRNA in the same sample. Concentrations of mRNA for carcinoembryonic antigen (CEA) were determined by using a qRT–PCR constructed in the laboratory using the TaqMan EZ technology (Applied Biosystems) and an external RNA copy standard (Öberg et al, 2004 (link)). CEA mRNA levels were estimated by dividing the CEA mRNA concentration with the concentration of 18S rRNA in the same sample, as determined by qRT–PCR for 18S rRNA (Applied Biosystems) and an external standard of total RNA from polyclonally activated human PBMCs. An amount of 1 pg RNA is defined as one unit (U) of 18S rRNA that corresponds approximately to 1 epithelial cell (Fahlgren et al, 2003 (link)).