TransplantImmunology 1999; 7:127-128 ARMS-PCR methodologies to determine IL-10, TNF.- , TNF-13 and TGF-131 gene polymo hlsms Chris Perrey a, Stephen J Turner b, Vera Pravica a, W Martin Howell b and Ian V Hutchinson a aSchool of Biological Sciences, University of Manchester and bHistocompatibility and Immunogenetics Laboratory, Southampton General Hospital Recently, we have described methods established to type poly- morphisms in interleuldn (IL)-10, turnout necrosis factor (TNF)-a, transforming growth factor (TGF)-[tl and interferon (IFN)-T genes, which have been shown to be associated with high or low expression levels of the cytokine in question. 1-4 These involved the polymerase chain reaction (PCR) and either sequence-specific oligonucleotide probing (SSOP) or the determination of microsatellite alleles by gel electrophoresis. 5 However, the PCR-SSOP approach is inappropriate for typing individual samples, so we developed amplification refractory mutation system (ARMS)-PCR methodologies for genotyping previously described IL-10, TNF-a and TGF-[ll gene poly- morphisms. In addition, we have extended this method for typing the TNF-13 Intron 1 Ncol restriction polymorphism, which has also been reported to be associated with high or low levels of both TNF-a and TNF-[! expression. 6'7 ARMS-PCR reaction mixes and PCR conditions are derived from those used in one of our laboratories for human leucocyte antigen genotyping, s The primer sequences and conditions are as follows: IL-10(-1082) IL-10(-1082) IL-10(-1082) (PCR product size = 258 bp) Generic primer (antisense): Primer G (sense): Primer A (sense): IL-10 (-8191/-592)* IL-10 (-819/-592)* IL-10 (-819/-592)* (PCR product size = 233 bp) Generic primer (antisense): Primer C (sense): Primer T (sense): TNF-a -308 TNF-a -308 TNF-a -308 (PCR product size = 184 bp) Generic primer (antisense): Primer G (sense): Primer A (sense): TGF-~I Codon 10 TGF-~I Codon 10 TGF-~z Codon 10 (PCR product size = 241 bp) Generic primer (sense): Primer C (antisense): Primer T (antisense): Address for correspondence:C Perrey, School of Biological Sciences, Universityof Manchester, Oxford Road, Manchester M13 9P~, UIC 5"-cagtgccaactgagaatttgg-Y 5"-ctactaaggcttctttgggag-3' 5"-actactaaggcttctttgggaa-Y 5"-aggatgtgttccaggctcct-3' 5"-cccttgtacaggtgatgtaac-3' 5"-acccffgtacaggtgatgtaat-Y 5"-tctcggtttcttctccatcg-3" 5'-ataggttttgaggggcatgg-Y (TNFI allele) 5"-aataggttttgaggggcatga-3' (12qF2 allele) 5'-tccgtgggatactgagacac-3" 5"-gcagcggtagcagcagcg-3" 5'-agcagcggtagcagcagca-3" ©Arnold 1999 0966-3274(99)TI265XX