Linked Life Data

9673693

Source:http://linkedlifedata.com/resource/pubmed/id/9673693

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
4
pubmed:dateCreated
1998-11-12
pubmed:abstractText
It is well established that nutrition is an important regulator of both serum insulin-like growth factor-I (IGF-1) and its binding proteins (IGFBPs). The Western ligand blot method (WLB) for simultaneous determinations of IGFBPs in serum or plasma samples was evaluated and validated with emphasis on its reproducible capabilities. After electrophoretic separation and transfer, the membranes were incubated with a mixture of recombinant labeled human (GF-I/IGF-II(rhIGF-I/rhlGF-II) and band intensities measured by autoradiography. The typical electrophoretic profile for pig serum, as determined with molecular weight markers, showed four mainbands of approximately 42-39, 32, 30-28 and 24 kDa which seemed to correspond to IGFBP-3, IGFBP-2, IGFBP-1 and IGFBP-4 respectively. Likewise, a triplet of approximately 42-39 kDa (IGFBP-3), a broad area called IGFBP-30 region (most probably IGFBP-1, -2 and -3 variants) and a third band of approximately 24 kDa (IGFBP-4) were seen in rat samples. Determination of IGFBP-2 and -1 in rat serum samples, as two separate bands on 12% gels was difficult due to their close electrophoretic migration and possibly to the reported lower levels of IGFBP-2 in adult rat serum. Dilutions tested on 0.2 micron nitrocellulose membranes with samples volumes between 0.25 to 1.5 microliters (1:10-1:60 dilutions), showed IGFBPs curves with good linearity which suggest first, that there exist a quantitative relation between the amount of each protein and the densitometric response and second, that the transfer of the proteins was linear across the range of 0.25 to 1.5 microliters (1:10-1:60 dilutions). Moreover, the results also suggest that losses were equally spread and that the proteins retained their binding properties after the transfer process. Reproducibility showed intraassay coefficients of variation (CVs) of 15% or lower using either a transfer device without cooling system or a combination of a transfer device with cooling system and manually defined band boundaries. In summary, it was shown that the optimized experimental conditions here described for the WLB method, allow reliable simultaneous measurements of the main pig and rat serum IGFBPs and therefore, could be utilised to detect changes in the serum profile after dietary manipulations.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Dec
pubmed:issn
0004-0622
pubmed:author
pubmed:issnType
Print
pubmed:volume
47
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
331-7
pubmed:dateRevised
2009-11-11
pubmed:meshHeading
pubmed:year
1997
pubmed:articleTitle
Reliability of the western ligand blot method for the simultaneous relative estimations of serum insulin-like growth factor binding proteins (IGFBPs).
pubmed:affiliation
Department of Chemistry, Universidad Nacional de Colombia, Santa Fe de Bogotá, Colombia.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't