Linked Life Data

2140249

Source:http://linkedlifedata.com/resource/pubmed/id/2140249

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
1990-6-21
pubmed:abstractText
A chemiluminescent reaction based on the deprotection of a phosphorylated phenyl dioxetane by alkaline phosphatase has recently been described (Schaap, A.P., 1988, J. Biolumin. Chemilumin. 2, 253). Light output is enhanced by intermolecular energy transfer to a micelle-solubilized fluorophore. This system is applied here to the detection of DNA probes on Southern blots. Enzyme solution assays which give an indication of sensitivity show that using this substrate 100 fg (0.7 amol) alkaline phosphatase can be detected on a luminescence plate reader (200 ms reading time). In a model Southern blotting system 180 fg HindIII digested lambda DNA was detected on film with homologous biotinylated DNA and a streptavidin-alkaline phosphatase complex. The single copy genes mos and raf-1, representing targets of 4.2 and 2.4 pg target DNA respectively, have also been detected in Southern-blotted human genomic DNA. A delay in reaching a plateau level of light output which is dependent on pH is observed but signal continues for at least 7 days. Typically, 12-h exposures to X-ray film were performed but once a steady-state light output had been achieved this time could be reduced to 2 h by preflashing film. This detection system represents a sensitive nonradioactive method, which is applicable not only to Southern blots but also to Northern and Western blots and any assay in which alkaline phosphatase is the label.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
0003-2697
pubmed:author
pubmed:issnType
Print
pubmed:volume
185
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
353-8
pubmed:dateRevised
2004-11-17
pubmed:meshHeading
pubmed:year
1990
pubmed:articleTitle
Nonradioactive DNA detection on Southern blots by enzymatically triggered chemiluminescence.
pubmed:affiliation
Corporate Research, Amersham International plc, Pollards Wood Laboratories, Bucks, United Kingdom.
pubmed:publicationType
Journal Article