Isolation of immunoglobulins and immunoaffinity HPLC

Josić, Dj.; Hofmann, Werner; Habermann, R.; Reutter, Werner

doi:10.1007/bf02316453

Cited by 2 publications

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“…The monospecific polyclonal anti-CBP65/67 antibodies were used for the immunoreaction as reported elsewhere (JosiC et al, 1988). Visualization of immunoreaction was performed with the chemiluminescent reagent, luminol.…”

Section: Methodsmentioning

confidence: 99%

“…Aliquots of each solubilized organ were analysed by SDSPAGE (Laemmli et al, 1970), stained with Coomassie brilliant blue R-250 or detected by Western blot according to Maniatis et al (1987). The monospecific polyclonal anti-CBP65/67 antibodies were used for the immunoreaction as reported elsewhere (JosiC et al, 1988). Visualization of immunoreaction was performed with the chemiluminescent reagent, luminol.…”

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confidence: 99%

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cDNA Cloning and Tissue‐specific Regulation of Expression of Rat Calcium‐binding Protein 65/67

Fan

Josić

Lim

et al. 1995

European Journal of Biochemistry

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We isolated a cDNA encoding the rat membrane-associated 65167-kDa calcium-binding protein, CBP 65/67, from a A ZAP I1 cDNA-expression library of rat liver by immunoscreening using monospecific polyclonal anti-(CBP 65/67) antibodies and monoclonal anti-(CBP 65/67) IgG. The product of this cDNA expressed in Escherichia coli was confirmed as CBP 65167 both by immunostaining and by comparison of the molecular mass with the CBP 65/67 isolated from rat liver by SDSPAGE. The cDNA sequence and the deduced amino acid sequence of CBP 65/67 both show a high degree of identity to human p68 and human calelectrin, which belong to a family of calcium-dependent, membrane-associated, phospholipid-binding proteins, called annexins. This means that CBP 65/67 is a homolog of the two human proteins just mentioned above. We are not aware that a rat annexin VI has previously been isolated and sequenced.The mRNA expression of CBP 65/67 in different rat organs during development was investigated by Northern blot analysis. In adult tissues, high mRNA levels of CBP 65/67 were found in lung, heart, muscle, spleen and especially in thymus and pancreas, whereas in liver, kidney, intestine, stomach and brain only low levels of CBP 65/67 mRNA could be detected. The amount of mRNA during tissue development in kidney, stomach and muscle showed only slight changes. In contrast, a significant increase of CBP 65/67 expression was observed in liver, lung, heart and brain. In most of the organs investigated, the level of mRNA correlated closely with the level of protein expression, indicating that the expression of CBP 65/67 in most organs is controlled primarily at the transcriptional level.Keywords. Calcium-binding protein; annexin VI; tissue-specific expression; expression regulation.Annexins are a family of Ca2+-dependent, membrane-associated, phospholipid-binding proteins (Crumpton and Dedman, 1990). More than ten different annexins, variously designated as lipocortins (Pepinsky et al., 1988;Gerke and Weber, 1984;Gerke, 1990;Browning et al., 1990), protein I, I1 and 111 (Shadle et al., 19851, calpactins (Saris et al., 1986), pp4-X (Romisch andHeimburger, 1990) endonexins I and I1 (Siidhof et al., 1983(Siidhof et al., , 1984, calelectrin (Geisow et al., 1986; Siidhof et al., 1988), lymphocyte Ca2+-binding protein p68 (Crompton et al., 1988), anchorin cII, synexin (Burns et al., 1989), calcimedins and placental anticoagulant proteins (Funakoski et al., 1987) have now been isolated and Characterized. The annexin family is widely distributed in nature: members of this protein family have been identified not only in mammals but also in different lower phyla, e.g. Drosophila (Johnston et al., 1990), Hydra (Schlaepfer et al., 1991), Dictyosteliurn (Doring et al., 1991), as well as in higher (EC 3.4.21.19). Note. The novel nucleotide sequence data published here have been deposited with the EMBL sequence data bank and are available under the accession number X86086. The novel amino acid sequence data have also been deposited with the EMBL sequence...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

cDNA Cloning and Tissue‐specific Regulation of Expression of Rat Calcium‐binding Protein 65/67

Fan

Josić

Lim

et al. 1995

European Journal of Biochemistry

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“…Trapping and purification 391 are often done by the aid of mixed-bed ion-exchange, hydroxyapatite, or HIC, or by some of the IgG-specific affinity schemes described later in this section. 392 Characteristic of polyclonal antibodies is a large heterogeneity in binding strength to the antigen, often to different regions (epitopes) of the antigen. This property can make polyclonal antibodies valuable in enrichment schemes, as exemplified by the on-line extraction of the benzodiazepine flunitrazepam and its main metabolites, 393 and isolation of aflatoxins, 394 but they are less useful as mediators of analytical separation due to the substantial variation in binding strengths towards single antigens or haptens.…”

Section: Bioaffinity Chromatographymentioning

confidence: 99%

High‐Performance Liquid Chromatography of Biological Macromolecules

Irgum¹

2000

Encyclopedia of Analytical Chemistry

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Isolation of immunoglobulins and immunoaffinity HPLC

Cited by 2 publications

References 1 publication

cDNA Cloning and Tissue‐specific Regulation of Expression of Rat Calcium‐binding Protein 65/67

cDNA Cloning and Tissue‐specific Regulation of Expression of Rat Calcium‐binding Protein 65/67

High‐Performance Liquid Chromatography of Biological Macromolecules

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