Miltenyi Biotec), mouse anti-CD11c APC(17-0114, eBioscience), mouse anti-CD45 vioblue (130-102-430, Miltenyi Biotec), mouse nti-CD3 Violetfluor 450 (75-0032, Tonbo biosciences), mouse anti-Prominin1 APC (130-092-335, Miltenyi Biotec) Results are expressed as ratio of immune cells/tubular cells, an internal control allowing to quantify immune cells infiltrate in the whole kidney, pp.130-102 ,
Qiagen) according to manufacturer's instructions. RNA concentration was measured by using NanoDrop1000 spectrophotometer (ThermoScientific) RNA were reverse transcribed using Maxima First Strand cDNA Synthesis Kit (Thermoscientific) and PCR was performed using SYBR green and specific primers on a light cycler 480 (Roche) Expression levels were normalized to the house keeping gene GusB (beta-glucuronidase) or 18 S using lightcycler advanced relative quantification program (Roche) Primers used were from eurogentec: Rabbit Cast s: AGCCAGCAAGTCGCTCAG and as: CCATCTCTTTGCTGATTGGAA, mouse Cast s: TCGCAAGTT GGTGGTACAAG and as: CTCCCCAAACTTGCTGCTT, mouse Calpn1 s: AGTGGAAAGGACCCTGGAGT and as: TCTCGTTCATAGGGGTCCAC, mouse Calpn2 s: TGGCTTCGGCATCTATGAG and as: AAGTTTTTGCC GAGGTGGAT, mouse IL1 alpha s: TTGGTTAAATGACCTGCAACA and as: GAGCGCTCACGAACAGTTG, mouse IL1 beta s: TGTAATGAAAGACGGCACACC and as: TCTTCTTTGGGTATTGCTTGG, mouse p21 var1, mRNA from kidney cortex was extracted using the RNeasy kit s: TGCGCTTGGAGTGATAGAAA and as: AACATCTCAGGGCCGAAA, mouse Gusb s: CTCTGGTGGCCTTAC CTGAT and as: CAGTTGTTGTCACCTTCACCTC, mouse 18 S s: AAGCATTCTGAAATTGGCTCA and as ,
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