By Robert K. Ockner (auth.), Jan F. C. Glatz, Ger J. Van Der Vusse (eds.)
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Molecular and Cellular Biochemistry 98: 27-33, 1990. © 1990 Kluwer Academic Publishers. Localization of liver fatty acid-binding protein and its mRNA in the liver and jejunum of rats: an immunohistochemical and in situ hybridization study Shoichi Isekil, Hisatake Kond02 , Masahiro HitomP and Teruo On0 4 1, 2 Department of Anatomy, Kanazawa University School of Medicine, Kanazawa, 920 Japan; 3,4 Department of Biochemistry, Niigata University School of Medicine, Niigata, 951 Japan Key words: liver fatty acid-binding protein, immunohistochemistry, in situ hybridization, liver, jejunum, rat Summary The localization of liver fatty acid-binding protein (L-FABP) and its mRNA in the liver and jejunum was examined in normal and 3-day-fasted rats by means of immunohistochemistry using a specific antibody to L-FABP and in situ hybridization using a synthetic oligonucleotide complementary to L-FABP mRNA as probe.
In situ hybridization The oligonucleotide probe was labeled by 3' -tailing to a specific activity of 1-2 X 108 cpmlJLg using [a35S]-dCTP (New England Nuclear). The hybridization procedure was essentially as described previously [17,18]. 015M sodium citrate), 1 X Denhardt's solution, 2% Sarkosyl,. 2), and then with a hybridization mixture containing 5 X 105 dpm of the labeled L-FABP probe. For the methodological control experiment, an excess amount (50 times that of the labeled probe) of the unlabeled template oligonucleotide complementary to the probe was added to the mixture.
1 % trifluoroacetic acid in acetonitrile were pooled. Peptides SP1 and SP2 were conjugated to Keyhole Limpet Hemocyanine (KLH) with maleimidobenzoyl-N-hydroxysuccinimide  using the cysteine-SH groups as described elsewhere . The peptides SP3 and SP4 were coupled to KLH using a glutaraldehyde coupling method . Control conjugates of the peptides with bovine serum albumin were produced using a carbodiimide coupling . Isolation and purification of FABPc Human H-FABPc was isolated from tissue obtained within 12 hours post mortem.