Semiquantitation of human chemokine mRNA levels with a newly constructed multispecific competitor fragment.
Chemokines are a group of inducible, locally acting proinflammatory cytokines which have been implicated in the pathogenesis of a variety of diseases. Important members of the group include monocyte chemoattractant protein (MCP)-1, -2, -3, macrophage inhibitory protein (MIP)-1alpha, -1beta and RANTES (regulated upon activation, normal T expressed and secreted). To facilitate further investigation of the human chemokines, we have constructed a novel multispecific competitor fragment containing primer binding sites for the CC-chemokines MCP-1, MCP-2, MCP-3, MIP-1alpha, MIP-1beta and RANTES, the CXC-chemokines MIP-2alpha, MIP-2beta as well as for the housekeeping gene beta-actin. Using this competitor fragment we can demonstrate reliable semiquantitation of reverse transcribed chemokine mRNAs. The assay should be useful for further studies, in particular for the semiquantitation of chemokine mRNA species from small cell or tissue specimens.
Dumoulin
F L
FL
Department of General Internal Medicine, University of Bonn, Germany. dumoulin@uni-bonn.de
Altfeld
M
M
Rockstroh
J K
JK
Leifeld
L
L
Sauerbruch
T
T
Spengler
U
U
D000199:Actins / Q000235:genetics; D001665:Binding Sites; D001667:Binding, Competitive; D018925:Chemokines / Q000235:genetics*; D017931:DNA Primers; D005784:Gene Amplification; D006801:Humans; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis*; D014158:Transcription, Genetic; D020298:U937 Cells
D000199:Actins; D018925:Chemokines; D017931:DNA Primers; D008070:Lipopolysaccharides; D012333:RNA, Messenger