A New Principle for Measurement of Cobalamin and Corrinoids, Used for Studies of Cobalamin Analogs on Serum Haptocorrin
Transcobalamin (TC) and haptocorrin (HC) are serum corrinoid-binding proteins. We developed new methods for measurement of the corrinoids bound to HC and TC. TC (n = 10) or HC (n = 138) was immunoprecipitated, and corrinoids were released by enzymatic degradation [subtilisin Carlsberg (EC 3.4.21.62)...
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| Veröffentlicht in: | Clinical chemistry (Baltimore, Md.) Md.), 2009-05, Vol.55 (5), p.1002-1010 |
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| creator | Hardlei, Tore Forsingdal Nexo, Ebba |
| description | Transcobalamin (TC) and haptocorrin (HC) are serum corrinoid-binding proteins. We developed new methods for measurement of the corrinoids bound to HC and TC.
TC (n = 10) or HC (n = 138) was immunoprecipitated, and corrinoids were released by enzymatic degradation [subtilisin Carlsberg (EC 3.4.21.62)] of the binding proteins. Binding of the released corrinoids to added unsaturated TC (apoTC) or HC (apoHC) created holoTC (as measure of cobalamins) and holoHC (as measure of corrinoids). holoTC and holoHC were measured by use of ELISA. The amounts of analogs were calculated as the difference between corrinoids and cobalamins. Corrinoids extracted from HC were separated with HPLC after addition of potassium cyanide (n = 3).
The corrinoid- and cobalamin-specific assays had a positive linear relation between analyte concentration and assay signal, detection limits of 8 and 4 pmol/L, and imprecision values (CV) of |
| doi_str_mv | 10.1373/clinchem.2008.114132 |
| format | Article |
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TC (n = 10) or HC (n = 138) was immunoprecipitated, and corrinoids were released by enzymatic degradation [subtilisin Carlsberg (EC 3.4.21.62)] of the binding proteins. Binding of the released corrinoids to added unsaturated TC (apoTC) or HC (apoHC) created holoTC (as measure of cobalamins) and holoHC (as measure of corrinoids). holoTC and holoHC were measured by use of ELISA. The amounts of analogs were calculated as the difference between corrinoids and cobalamins. Corrinoids extracted from HC were separated with HPLC after addition of potassium cyanide (n = 3).
The corrinoid- and cobalamin-specific assays had a positive linear relation between analyte concentration and assay signal, detection limits of 8 and 4 pmol/L, and imprecision values (CV) of <or=10% and <or=13% for concentrations between 45-200 and 12-115 pmol/L, respectively. No analogs were bound to serum TC, whereas the mean (95% reference range) for analogs present on HC was 245 (100-380) pmol/L. On HPLC a substantial amount of the analogs showed elution patterns similar to those of dicyanocobinamide.
Our methods for measurement of unmodified corrinoids in serum demonstrate that HC carries cobalamin analogs not recognized by TC, and that on HPLC a substantial part of these analogs elute similarly to cobinamide.</description><identifier>ISSN: 0009-9147</identifier><identifier>EISSN: 1530-8561</identifier><identifier>DOI: 10.1373/clinchem.2008.114132</identifier><identifier>PMID: 19299543</identifier><identifier>CODEN: CLCHAU</identifier><language>eng</language><publisher>Washington, DC: Am Assoc Clin Chem</publisher><subject>Adult ; Analytical, structural and metabolic biochemistry ; Biological and medical sciences ; Biomedical research ; Blood & organ donations ; Blood banks ; Chromatography, High Pressure Liquid - methods ; Corrinoids - blood ; Detection limits ; Enzyme-Linked Immunosorbent Assay - methods ; Female ; Fluorides ; Fundamental and applied biological sciences. Psychology ; Humans ; Immunoprecipitation ; Investigative techniques, diagnostic techniques (general aspects) ; Linear Models ; Liquid chromatography ; Male ; Medical sciences ; Middle Aged ; Protein Binding ; Proteins ; Studies ; Transcobalamins - metabolism ; Vitamin B 12 - blood ; Young Adult</subject><ispartof>Clinical chemistry (Baltimore, Md.), 2009-05, Vol.55 (5), p.1002-1010</ispartof><rights>2009 INIST-CNRS</rights><rights>Copyright American Association for Clinical Chemistry May 2009</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c439t-235148481e762abf06b40632ad92e3b61ff970320c79e1d9f7a51cd5b432b1993</citedby><cites>FETCH-LOGICAL-c439t-235148481e762abf06b40632ad92e3b61ff970320c79e1d9f7a51cd5b432b1993</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>315,781,785,27926,27927</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=21459495$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/19299543$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Hardlei, Tore Forsingdal</creatorcontrib><creatorcontrib>Nexo, Ebba</creatorcontrib><title>A New Principle for Measurement of Cobalamin and Corrinoids, Used for Studies of Cobalamin Analogs on Serum Haptocorrin</title><title>Clinical chemistry (Baltimore, Md.)</title><addtitle>Clin Chem</addtitle><description>Transcobalamin (TC) and haptocorrin (HC) are serum corrinoid-binding proteins. We developed new methods for measurement of the corrinoids bound to HC and TC.
TC (n = 10) or HC (n = 138) was immunoprecipitated, and corrinoids were released by enzymatic degradation [subtilisin Carlsberg (EC 3.4.21.62)] of the binding proteins. Binding of the released corrinoids to added unsaturated TC (apoTC) or HC (apoHC) created holoTC (as measure of cobalamins) and holoHC (as measure of corrinoids). holoTC and holoHC were measured by use of ELISA. The amounts of analogs were calculated as the difference between corrinoids and cobalamins. Corrinoids extracted from HC were separated with HPLC after addition of potassium cyanide (n = 3).
The corrinoid- and cobalamin-specific assays had a positive linear relation between analyte concentration and assay signal, detection limits of 8 and 4 pmol/L, and imprecision values (CV) of <or=10% and <or=13% for concentrations between 45-200 and 12-115 pmol/L, respectively. No analogs were bound to serum TC, whereas the mean (95% reference range) for analogs present on HC was 245 (100-380) pmol/L. On HPLC a substantial amount of the analogs showed elution patterns similar to those of dicyanocobinamide.
Our methods for measurement of unmodified corrinoids in serum demonstrate that HC carries cobalamin analogs not recognized by TC, and that on HPLC a substantial part of these analogs elute similarly to cobinamide.</description><subject>Adult</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Biological and medical sciences</subject><subject>Biomedical research</subject><subject>Blood & organ donations</subject><subject>Blood banks</subject><subject>Chromatography, High Pressure Liquid - methods</subject><subject>Corrinoids - blood</subject><subject>Detection limits</subject><subject>Enzyme-Linked Immunosorbent Assay - methods</subject><subject>Female</subject><subject>Fluorides</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Humans</subject><subject>Immunoprecipitation</subject><subject>Investigative techniques, diagnostic techniques (general aspects)</subject><subject>Linear Models</subject><subject>Liquid chromatography</subject><subject>Male</subject><subject>Medical sciences</subject><subject>Middle Aged</subject><subject>Protein Binding</subject><subject>Proteins</subject><subject>Studies</subject><subject>Transcobalamins - metabolism</subject><subject>Vitamin B 12 - blood</subject><subject>Young Adult</subject><issn>0009-9147</issn><issn>1530-8561</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNpdkV1rFDEUhoModq3-A5Eg6JWz5uRjZnK5LGoLtRVqr0Nm5kw3JTNZkxkG_71pd7XUq3DC876cw0PIW2BrEJX43Ho3tjsc1pyxeg0gQfBnZAVKsKJWJTwnK8aYLjTI6oS8Sukuj7Kqy5fkBDTXWkmxIsuGXuJCf8Rc5vYeaR8i_Y42zREHHCcaeroNjfV2cCO1Y5enmOHguvSJ3iTsHhLX09w5TE_pzWh9uM2fI73GOA_0zO6n0D7kX5MXvfUJ3xzfU3Lz9cvP7VlxcfXtfLu5KFop9FRwoUDWsgasSm6bnpWNZKXgttMcRVNC3-uKCc7aSiN0uq-sgrZTjRS8Aa3FKfl46N3H8GvGNJnBpRa9tyOGOZmyglKIimfw_X_gXZhjviAZDkLXUgrIkDxAbQwpRezNPrrBxt8GmLm3Yv5aMfdWzMFKjr07ds_NgN1j6KghAx-OgE2t9X202Ub6x3GQSkutHq_Zudvd4iKaNFjvcy2YZVmUMipvwrj4AyPZo54</recordid><startdate>20090501</startdate><enddate>20090501</enddate><creator>Hardlei, Tore Forsingdal</creator><creator>Nexo, Ebba</creator><general>Am Assoc Clin Chem</general><general>American Association for Clinical Chemistry</general><general>Oxford University Press</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>4U-</scope><scope>7QO</scope><scope>7RV</scope><scope>7TM</scope><scope>7U7</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>88I</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>BKSAR</scope><scope>C1K</scope><scope>CCPQU</scope><scope>D1I</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>KB.</scope><scope>KB0</scope><scope>M0S</scope><scope>M1P</scope><scope>M2P</scope><scope>NAPCQ</scope><scope>P64</scope><scope>PCBAR</scope><scope>PDBOC</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>Q9U</scope><scope>RC3</scope><scope>S0X</scope><scope>7X8</scope></search><sort><creationdate>20090501</creationdate><title>A New Principle for Measurement of Cobalamin and Corrinoids, Used for Studies of Cobalamin Analogs on Serum Haptocorrin</title><author>Hardlei, Tore Forsingdal ; Nexo, Ebba</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c439t-235148481e762abf06b40632ad92e3b61ff970320c79e1d9f7a51cd5b432b1993</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2009</creationdate><topic>Adult</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Biological and medical sciences</topic><topic>Biomedical research</topic><topic>Blood & organ donations</topic><topic>Blood banks</topic><topic>Chromatography, High Pressure Liquid - methods</topic><topic>Corrinoids - blood</topic><topic>Detection limits</topic><topic>Enzyme-Linked Immunosorbent Assay - methods</topic><topic>Female</topic><topic>Fluorides</topic><topic>Fundamental and applied biological sciences. 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We developed new methods for measurement of the corrinoids bound to HC and TC.
TC (n = 10) or HC (n = 138) was immunoprecipitated, and corrinoids were released by enzymatic degradation [subtilisin Carlsberg (EC 3.4.21.62)] of the binding proteins. Binding of the released corrinoids to added unsaturated TC (apoTC) or HC (apoHC) created holoTC (as measure of cobalamins) and holoHC (as measure of corrinoids). holoTC and holoHC were measured by use of ELISA. The amounts of analogs were calculated as the difference between corrinoids and cobalamins. Corrinoids extracted from HC were separated with HPLC after addition of potassium cyanide (n = 3).
The corrinoid- and cobalamin-specific assays had a positive linear relation between analyte concentration and assay signal, detection limits of 8 and 4 pmol/L, and imprecision values (CV) of <or=10% and <or=13% for concentrations between 45-200 and 12-115 pmol/L, respectively. No analogs were bound to serum TC, whereas the mean (95% reference range) for analogs present on HC was 245 (100-380) pmol/L. On HPLC a substantial amount of the analogs showed elution patterns similar to those of dicyanocobinamide.
Our methods for measurement of unmodified corrinoids in serum demonstrate that HC carries cobalamin analogs not recognized by TC, and that on HPLC a substantial part of these analogs elute similarly to cobinamide.</abstract><cop>Washington, DC</cop><pub>Am Assoc Clin Chem</pub><pmid>19299543</pmid><doi>10.1373/clinchem.2008.114132</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record> |
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| identifier | ISSN: 0009-9147 |
| ispartof | Clinical chemistry (Baltimore, Md.), 2009-05, Vol.55 (5), p.1002-1010 |
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| language | eng |
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| subjects | Adult Analytical, structural and metabolic biochemistry Biological and medical sciences Biomedical research Blood & organ donations Blood banks Chromatography, High Pressure Liquid - methods Corrinoids - blood Detection limits Enzyme-Linked Immunosorbent Assay - methods Female Fluorides Fundamental and applied biological sciences. Psychology Humans Immunoprecipitation Investigative techniques, diagnostic techniques (general aspects) Linear Models Liquid chromatography Male Medical sciences Middle Aged Protein Binding Proteins Studies Transcobalamins - metabolism Vitamin B 12 - blood Young Adult |
| title | A New Principle for Measurement of Cobalamin and Corrinoids, Used for Studies of Cobalamin Analogs on Serum Haptocorrin |
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