A simplified method for assessing PHA induced stimulation of rat peripheral blood lymphocytes

Keller, Steven E.; Schleifer, Steven J.; McKegney, F. Patrick; Sherman, Jeffrey; Camerino, Maria; Stein, Mary Kay

doi:10.1016/0022-1759(82)90395-7

Cited by 20 publications

(4 citation statements)

References 11 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Aliquots (1 pg protein) of pure CNS or PNS myelin were suspended in 25 ~1 of 50 mM Na borate buffer (pH 8) and frozen (-80°C) until used for iodination. Circulating lymphocytes extracted from blood (Keller et al, 1982) were aliquoted, pelleted, and frozen (-SOY) until used. Thawed lymphocytes (30 fig protein) were resuspended in 50 mM Na borate buffer, sdnicated on ice, and the membranes pelleted (100.000 x P for 10 min).…”

Section: Methodsmentioning

confidence: 99%

Molecular cloning of a 2',3'-cyclic nucleotide 3'-phosphodiesterase: mRNAs with different 5' ends encode the same set of proteins in nervous and lymphoid tissues

Bernier¹,

Álvarez²,

Em³

et al. 1987

J. Neurosci.

137

View full text Add to dashboard Cite

Antibodies raised to a mixture of the 46 and 48 kDa rat CNS 2',3'-cyclic nucleotide 3-phosphodiesterases (CNPs) recognized apparently identical proteins in peripheral nervous system (PNS), thymus, and circulating blood lymphocytes. These antibodies were used to identify, in a rat brain phage lambda gt11 expression library, cDNA clones encoding beta-galactosidase-CNP fusion proteins, some of which showed CNP activity. In RNA blots, the subcloned CNP cDNA inserts hybridized to mRNAs of approximately 2400 and approximately 2800 nucleotides (nts), and to a approximately 2500 nt mRNA from thymus. Several nonexpressing CNP cDNAs were identified by plaque hybridization, and the mRNA transcribed in vitro from one of these cDNAs (pCNP7) encoded a complete 46 kDa CNP polypeptide. Examination of the deduced amino acid sequence revealed an apparent homology to cAMP binding sites in several other proteins. A 373 bp segment from the 5' end of this pCNP7 hybridized only to the 2800 nt nervous system mRNAs, thus revealing that not all CNP mRNAs share the same 5'-ends. Genomic DNA blots probed with CNP cDNAs suggest that there is a single gene which can be alternatively spliced to produce the various mRNA transcripts in the nervous and lymphoid tissues.

show abstract

Section: Methodsmentioning

confidence: 99%

Molecular cloning of a 2',3'-cyclic nucleotide 3'-phosphodiesterase: mRNAs with different 5' ends encode the same set of proteins in nervous and lymphoid tissues

Bernier¹,

Álvarez²,

Em³

et al. 1987

J. Neurosci.

137

View full text Add to dashboard Cite

show abstract

“…The mixture was gently layered over Biocoll separation medium in a centrifuge tube. It was centrifuged at 400 × g for 40 min at 4 • C [16,17]. The clear lymphocyte layer at the interphase was collected using a syringe and washed with equal volume of PBS by spinning the suspension at 250 × g for 10 min at 4 • C. The pellet was homogeneously suspended in 1 ml of PBS.…”

Section: Preparation Of Whole Homogenates (Wh) Of Blood Lymphocytes (mentioning

confidence: 99%

In Vivo Exposure of Swiss Albino Mice to Chronic Low Dose of Dimethylnitrosamine (DMN) Lowers Poly-ADP-Ribosylation (PAR) of Bone Marrow Cell and Blood Lymphocyte Proteins

Kma

Sharan

2006

Mol Cell Biochem

View full text Add to dashboard Cite

Efforts to identify an easy and convenient biomarker of carcinogenesis with potentials of application in mass screening program continue. In a series of investigations on mice exposed to different carcinogens, poly-ADP-ribosylation (PAR) of cellular proteins of different tissues has been shown to be a potential biomarker of carcinogenesis. Because blood based biomarker of carcinogenesis offers significant advantage in its use in a cancer screening program, this investigation was undertaken to find correlations between initiation of carcinogenesis and PAR of bone marrow cell (BMC) and blood lymphocyte (BL) proteins in mice chronically exposed to low dose of dimethylnitrosamine (DMN) for up to four weeks in vivo. The exposure was either alone or in combination with 3-aminobenzamide (3-AB), an inhibitor of PAR. Total PAR of cellular proteins and of histone H1 protein were monitored by slot and Western blot immunoprobe assays, respectively. The PAR of total cellular proteins as well as of histone H1 was down-regulated in duration of exposure dependent manners. The results suggest that BMC and BL mirrored status of PAR in other tissues. This finding opens up the possibility of using PAR as a biomarker of carcinogenesis in a blood based test utilizing immunoprobe assay of cellular PAR.

show abstract

“…A splenocyte suspension was prepared by gentle homogenizing of the spleen with a glass homogenizer. Lymphocytes were isolated by 65 % Percoll (Pharmacia, Piscataway, N. J., USA) densitygradient sedimentation [20], washed four times in PBS, and finally suspended at 5 10 6 cells/ml in RPMI-1640 medium (Sigma) supplemented with 5 % heat-inactivated fetal calf serum (Gibco BRL, Grand Island, N. Y., USA). The proliferative response to a mitogen, ConA (Sigma), was measured as reported previously [21].…”

Section: Methodsmentioning

confidence: 99%

Proinsulin C peptide obviates sympathetically mediated suppression of splenic lymphocyte activity in rats

et al. 2000

View full text Add to dashboard Cite

A simplified method for assessing PHA induced stimulation of rat peripheral blood lymphocytes

Cited by 20 publications

References 11 publications

Molecular cloning of a 2',3'-cyclic nucleotide 3'-phosphodiesterase: mRNAs with different 5' ends encode the same set of proteins in nervous and lymphoid tissues

Molecular cloning of a 2',3'-cyclic nucleotide 3'-phosphodiesterase: mRNAs with different 5' ends encode the same set of proteins in nervous and lymphoid tissues

In Vivo Exposure of Swiss Albino Mice to Chronic Low Dose of Dimethylnitrosamine (DMN) Lowers Poly-ADP-Ribosylation (PAR) of Bone Marrow Cell and Blood Lymphocyte Proteins

Proinsulin C peptide obviates sympathetically mediated suppression of splenic lymphocyte activity in rats

Contact Info

Product

Resources

About