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Following differentiation at 28 days of culture, 1 × 10 differentiated cells were harvested after being detached with 0.25% trypsin (Sigma-Aldrich®), washed with PBS, permeabilized in 4% of paraformaldehyde, and incubated for 18 h at 4°C with the monoclonal antibodies at the same dilutions described for immunostaining in 1% Triton X-100 and 0.5% bovine serum albumin (BSA).
On day 7, 10 differentiated DC were cytospinned 5′ at 800 rpm.
One tumour group comprised 10 differentiated (classic) PTC of tumour stage pT1-pT4, according to the pTpNM system [ 21].
Approximately 1 × 10 differentiated cells in each well of 96-well plate were loaded with 10 μM Fluo-4 for 1 h at 37°C.
We found that the rectospheres formed tumor masses 2 weeks post injection, whereas 1 × 10 differentiated primary rectal cancer cells did not give rise to any detectable tumors at 8 weeks post injection.
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Histological classification was performed according to the World Health Organization (WHO) system in well-differentiated (G1; n = 39), moderately differentiated (G2; n = 32) and poorly differentiated (G3; n = 31) carcinomas.
The histopathologic types of these adenocarcinoma cases include 9 adenoma canceration (8.2%, 7 well-differentiated and 2 moderately differentiated), 29 well-differentiated adenocarcinoma (26.9%), 29 moderately differentiated adenocarcinomas (26.9%), 30 poorly differentiated adenocarcinoma (27.8%), and 11 mucinous adenocarcinomas (10.2%).
The tumour grading included 70 well differentiated (38.3%), 85 moderately differentiated (46.4%), and 28 poorly differentiated adenocarcinomas (15.3%).
Seventy-two cases were classified as SCC and 96 as adenocarcinoma (66 well differentiated, 80 moderately differentiated, and 22 poorly differentiated).
CFP-10 differentiated DCs (CFP10-DCs) are phenotypically and morphologically similar to DCs differentiated conventionally with GM-CSF [20].
Tumour differentiation was used as a binomial variable: 0=poorly or moderately differentiated and 1=well differentiated.
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