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Integrated single-label liquid-phase assay of APOE codons 112 and 158 and a lipoprotein study in British women

Integrated single-label liquid-phase assay of APOE codons 112 and 158 and a lipoprotein study in British women
Integrated single-label liquid-phase assay of APOE codons 112 and 158 and a lipoprotein study in British women
BACKGROUND: Apolipoprotein E (APOE) is an important element of lipid metabolism and, hence, cardiovascular disorders. APOE has 3 main allelic variants: epsilon3, epsilon4, and epsilon2. Of these, epsilon3 is the most common, followed by epsilon4 and epsilon2. The associations of these isoforms with cardiovascular disorders and Alzheimer disease have been widely studied in different populations. Most of the genotyping in these studies has been performed with gel-based methods, which have important limitations, particularly for large epidemiologic studies. We therefore developed an integrated "one-tube" liquid-phase assay. METHODS: To measure APOE isoforms, we developed an integrated single-label liquid-phase fluorescence assay containing 2 PCR primers, 2 probes, and 2 quencher oligonucleotides. We used a 384-well LightTyper, but the assay would be generically applicable for use with any fluorescence detector with thermal ramp control. We validated this method and applied it in the British Women's Heart and Health Study. RESULTS: There were 4 melting peaks, at 41, 56, 61, and 69 degrees C, which generated 6 distinctive patterns representing genotypic combinations of epsilon3, epsilon4, and epsilon2. The magnitude and direction of the associations found with total cholesterol, HDL-cholesterol, triglycerides, and estimated LDL-cholesterol were consistent with previous reports. CONCLUSION: The one-tube LightTyper assay presented here enables accurate, convenient, and economical genotyping of APOE and can be used for large epidemiologic studies
humans, blood, alzheimer disease, methods, fluorescence, population, ldl, triglycerides, research support, cholesterol, female, codon, great britain, epidemiologic studies, lipoproteins, report, research, apolipoproteins, apolipoproteins e, aged, disease, hdl, metabolism, lipid metabolism, laboratories, protein isoforms, middle aged, combination, epidemiology, heart, health, genotype, protein, genetics
1420-1423
Abdollahi, Mohammad Reza
96ec68db-6302-4216-9aaf-9951a44be8b8
Guthrie, Philip A.I.
2ad09527-826e-4d74-bb50-0962dacb0292
Smith, George Davey
f5bc8327-f2cb-49a0-8eae-4a6ba63207a2
Lawlor, Debbie A.
799826df-f115-4fb7-83ea-53c246c220d4
Ebrahim, Shah
0f2ade5c-4ef6-4ca7-9f9b-9b60ba192b13
Day, Ian N.M.
b749b30a-1f4c-40eb-af0e-a50427388b39
Abdollahi, Mohammad Reza
96ec68db-6302-4216-9aaf-9951a44be8b8
Guthrie, Philip A.I.
2ad09527-826e-4d74-bb50-0962dacb0292
Smith, George Davey
f5bc8327-f2cb-49a0-8eae-4a6ba63207a2
Lawlor, Debbie A.
799826df-f115-4fb7-83ea-53c246c220d4
Ebrahim, Shah
0f2ade5c-4ef6-4ca7-9f9b-9b60ba192b13
Day, Ian N.M.
b749b30a-1f4c-40eb-af0e-a50427388b39

Abdollahi, Mohammad Reza, Guthrie, Philip A.I., Smith, George Davey, Lawlor, Debbie A., Ebrahim, Shah and Day, Ian N.M. (2006) Integrated single-label liquid-phase assay of APOE codons 112 and 158 and a lipoprotein study in British women. Clinical Chemistry, 52 (7), 1420-1423. (doi:10.1373/clinchem.2006.067082). (PMID:16644874)

Record type: Article

Abstract

BACKGROUND: Apolipoprotein E (APOE) is an important element of lipid metabolism and, hence, cardiovascular disorders. APOE has 3 main allelic variants: epsilon3, epsilon4, and epsilon2. Of these, epsilon3 is the most common, followed by epsilon4 and epsilon2. The associations of these isoforms with cardiovascular disorders and Alzheimer disease have been widely studied in different populations. Most of the genotyping in these studies has been performed with gel-based methods, which have important limitations, particularly for large epidemiologic studies. We therefore developed an integrated "one-tube" liquid-phase assay. METHODS: To measure APOE isoforms, we developed an integrated single-label liquid-phase fluorescence assay containing 2 PCR primers, 2 probes, and 2 quencher oligonucleotides. We used a 384-well LightTyper, but the assay would be generically applicable for use with any fluorescence detector with thermal ramp control. We validated this method and applied it in the British Women's Heart and Health Study. RESULTS: There were 4 melting peaks, at 41, 56, 61, and 69 degrees C, which generated 6 distinctive patterns representing genotypic combinations of epsilon3, epsilon4, and epsilon2. The magnitude and direction of the associations found with total cholesterol, HDL-cholesterol, triglycerides, and estimated LDL-cholesterol were consistent with previous reports. CONCLUSION: The one-tube LightTyper assay presented here enables accurate, convenient, and economical genotyping of APOE and can be used for large epidemiologic studies

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More information

e-pub ahead of print date: 27 April 2006
Published date: July 2006
Keywords: humans, blood, alzheimer disease, methods, fluorescence, population, ldl, triglycerides, research support, cholesterol, female, codon, great britain, epidemiologic studies, lipoproteins, report, research, apolipoproteins, apolipoproteins e, aged, disease, hdl, metabolism, lipid metabolism, laboratories, protein isoforms, middle aged, combination, epidemiology, heart, health, genotype, protein, genetics
Organisations: Cancer Sciences

Identifiers

Local EPrints ID: 59443
URI: http://eprints.soton.ac.uk/id/eprint/59443
PURE UUID: 7170adc0-7e5a-4d53-9ba6-c2a302f7e5df

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Date deposited: 03 Sep 2008
Last modified: 13 Mar 2019 20:30

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