Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins
Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used...
Ausführliche Beschreibung
Gespeichert in:
| Autor*in: |
Finlay, W. J. J. [verfasserIn] DeVore, N. C. [verfasserIn] Dobrovolskaia, E. N. [verfasserIn] |
|---|
| Format: |
E-Artikel |
|---|
| Erschienen: |
Oxford, UK: Blackwell Science Ltd ; 2005 |
|---|
| Schlagwörter: |
|---|
| Umfang: |
Online-Ressource |
|---|
| Reproduktion: |
2005 ; Blackwell Publishing Journal Backfiles 1879-2005 |
|---|---|
| Übergeordnetes Werk: |
In: Clinical & experimental allergy - Oxford : Blackwell Science, 1989, 35(2005), 8, Seite 0 |
| Übergeordnetes Werk: |
volume:35 ; year:2005 ; number:8 ; pages:0 |
| Links: |
|---|
| DOI / URN: |
10.1111/j.1365-2222.2005.02307.x |
|---|
| Katalog-ID: |
NLEJ242601065 |
|---|
| LEADER | 01000caa a22002652 4500 | ||
|---|---|---|---|
| 001 | NLEJ242601065 | ||
| 003 | DE-627 | ||
| 005 | 20210707161450.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 120427s2005 xx |||||o 00| ||und c | ||
| 024 | 7 | |a 10.1111/j.1365-2222.2005.02307.x |2 doi | |
| 035 | |a (DE-627)NLEJ242601065 | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 100 | 1 | |a Finlay, W. J. J. |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins |
| 264 | 1 | |a Oxford, UK |b Blackwell Science Ltd |c 2005 | |
| 300 | |a Online-Ressource | ||
| 336 | |a nicht spezifiziert |b zzz |2 rdacontent | ||
| 337 | |a nicht spezifiziert |b z |2 rdamedia | ||
| 338 | |a nicht spezifiziert |b zu |2 rdacarrier | ||
| 520 | |a Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. | ||
| 533 | |d 2005 |f Blackwell Publishing Journal Backfiles 1879-2005 |7 |2005|||||||||| | ||
| 650 | 4 | |a 2D SDS-PAGE | |
| 700 | 1 | |a DeVore, N. C. |e verfasserin |4 aut | |
| 700 | 1 | |a Dobrovolskaia, E. N. |e verfasserin |4 aut | |
| 700 | 1 | |a Gam, A. |4 oth | |
| 700 | 1 | |a Goodyear, C. S. |4 oth | |
| 700 | 1 | |a Slater, J. E. |4 oth | |
| 773 | 0 | 8 | |i In |t Clinical & experimental allergy |d Oxford : Blackwell Science, 1989 |g 35(2005), 8, Seite 0 |h Online-Ressource |w (DE-627)NLEJ243926391 |w (DE-600)2004469-0 |x 1365-2222 |7 nnns |
| 773 | 1 | 8 | |g volume:35 |g year:2005 |g number:8 |g pages:0 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x |q text/html |x Verlag |z Deutschlandweit zugänglich |3 Volltext |
| 912 | |a GBV_USEFLAG_U | ||
| 912 | |a ZDB-1-DJB | ||
| 912 | |a GBV_NL_ARTICLE | ||
| 951 | |a AR | ||
| 952 | |d 35 |j 2005 |e 8 |h 0 | ||
| author_variant |
w j j f wjj wjjf n c d nc ncd e n d en end |
|---|---|
| matchkey_str |
article:13652222:2005----::xliighainmuolblnytmoipiyhgnrtoorcmiatni |
| hierarchy_sort_str |
2005 |
| publishDate |
2005 |
| allfields |
10.1111/j.1365-2222.2005.02307.x doi (DE-627)NLEJ242601065 DE-627 ger DE-627 rakwb Finlay, W. J. J. verfasserin aut Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins Oxford, UK Blackwell Science Ltd 2005 Online-Ressource nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. 2005 Blackwell Publishing Journal Backfiles 1879-2005 |2005|||||||||| 2D SDS-PAGE DeVore, N. C. verfasserin aut Dobrovolskaia, E. N. verfasserin aut Gam, A. oth Goodyear, C. S. oth Slater, J. E. oth In Clinical & experimental allergy Oxford : Blackwell Science, 1989 35(2005), 8, Seite 0 Online-Ressource (DE-627)NLEJ243926391 (DE-600)2004469-0 1365-2222 nnns volume:35 year:2005 number:8 pages:0 http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x text/html Verlag Deutschlandweit zugänglich Volltext GBV_USEFLAG_U ZDB-1-DJB GBV_NL_ARTICLE AR 35 2005 8 0 |
| spelling |
10.1111/j.1365-2222.2005.02307.x doi (DE-627)NLEJ242601065 DE-627 ger DE-627 rakwb Finlay, W. J. J. verfasserin aut Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins Oxford, UK Blackwell Science Ltd 2005 Online-Ressource nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. 2005 Blackwell Publishing Journal Backfiles 1879-2005 |2005|||||||||| 2D SDS-PAGE DeVore, N. C. verfasserin aut Dobrovolskaia, E. N. verfasserin aut Gam, A. oth Goodyear, C. S. oth Slater, J. E. oth In Clinical & experimental allergy Oxford : Blackwell Science, 1989 35(2005), 8, Seite 0 Online-Ressource (DE-627)NLEJ243926391 (DE-600)2004469-0 1365-2222 nnns volume:35 year:2005 number:8 pages:0 http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x text/html Verlag Deutschlandweit zugänglich Volltext GBV_USEFLAG_U ZDB-1-DJB GBV_NL_ARTICLE AR 35 2005 8 0 |
| allfields_unstemmed |
10.1111/j.1365-2222.2005.02307.x doi (DE-627)NLEJ242601065 DE-627 ger DE-627 rakwb Finlay, W. J. J. verfasserin aut Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins Oxford, UK Blackwell Science Ltd 2005 Online-Ressource nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. 2005 Blackwell Publishing Journal Backfiles 1879-2005 |2005|||||||||| 2D SDS-PAGE DeVore, N. C. verfasserin aut Dobrovolskaia, E. N. verfasserin aut Gam, A. oth Goodyear, C. S. oth Slater, J. E. oth In Clinical & experimental allergy Oxford : Blackwell Science, 1989 35(2005), 8, Seite 0 Online-Ressource (DE-627)NLEJ243926391 (DE-600)2004469-0 1365-2222 nnns volume:35 year:2005 number:8 pages:0 http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x text/html Verlag Deutschlandweit zugänglich Volltext GBV_USEFLAG_U ZDB-1-DJB GBV_NL_ARTICLE AR 35 2005 8 0 |
| allfieldsGer |
10.1111/j.1365-2222.2005.02307.x doi (DE-627)NLEJ242601065 DE-627 ger DE-627 rakwb Finlay, W. J. J. verfasserin aut Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins Oxford, UK Blackwell Science Ltd 2005 Online-Ressource nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. 2005 Blackwell Publishing Journal Backfiles 1879-2005 |2005|||||||||| 2D SDS-PAGE DeVore, N. C. verfasserin aut Dobrovolskaia, E. N. verfasserin aut Gam, A. oth Goodyear, C. S. oth Slater, J. E. oth In Clinical & experimental allergy Oxford : Blackwell Science, 1989 35(2005), 8, Seite 0 Online-Ressource (DE-627)NLEJ243926391 (DE-600)2004469-0 1365-2222 nnns volume:35 year:2005 number:8 pages:0 http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x text/html Verlag Deutschlandweit zugänglich Volltext GBV_USEFLAG_U ZDB-1-DJB GBV_NL_ARTICLE AR 35 2005 8 0 |
| allfieldsSound |
10.1111/j.1365-2222.2005.02307.x doi (DE-627)NLEJ242601065 DE-627 ger DE-627 rakwb Finlay, W. J. J. verfasserin aut Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins Oxford, UK Blackwell Science Ltd 2005 Online-Ressource nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. 2005 Blackwell Publishing Journal Backfiles 1879-2005 |2005|||||||||| 2D SDS-PAGE DeVore, N. C. verfasserin aut Dobrovolskaia, E. N. verfasserin aut Gam, A. oth Goodyear, C. S. oth Slater, J. E. oth In Clinical & experimental allergy Oxford : Blackwell Science, 1989 35(2005), 8, Seite 0 Online-Ressource (DE-627)NLEJ243926391 (DE-600)2004469-0 1365-2222 nnns volume:35 year:2005 number:8 pages:0 http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x text/html Verlag Deutschlandweit zugänglich Volltext GBV_USEFLAG_U ZDB-1-DJB GBV_NL_ARTICLE AR 35 2005 8 0 |
| source |
In Clinical & experimental allergy 35(2005), 8, Seite 0 volume:35 year:2005 number:8 pages:0 |
| sourceStr |
In Clinical & experimental allergy 35(2005), 8, Seite 0 volume:35 year:2005 number:8 pages:0 |
| format_phy_str_mv |
Article |
| institution |
findex.gbv.de |
| topic_facet |
2D SDS-PAGE |
| isfreeaccess_bool |
false |
| container_title |
Clinical & experimental allergy |
| authorswithroles_txt_mv |
Finlay, W. J. J. @@aut@@ DeVore, N. C. @@aut@@ Dobrovolskaia, E. N. @@aut@@ Gam, A. @@oth@@ Goodyear, C. S. @@oth@@ Slater, J. E. @@oth@@ |
| publishDateDaySort_date |
2005-01-01T00:00:00Z |
| hierarchy_top_id |
NLEJ243926391 |
| id |
NLEJ242601065 |
| fullrecord |
<?xml version="1.0" encoding="UTF-8"?><collection xmlns="http://www.loc.gov/MARC21/slim"><record><leader>01000caa a22002652 4500</leader><controlfield tag="001">NLEJ242601065</controlfield><controlfield tag="003">DE-627</controlfield><controlfield tag="005">20210707161450.0</controlfield><controlfield tag="007">cr uuu---uuuuu</controlfield><controlfield tag="008">120427s2005 xx |||||o 00| ||und c</controlfield><datafield tag="024" ind1="7" ind2=" "><subfield code="a">10.1111/j.1365-2222.2005.02307.x</subfield><subfield code="2">doi</subfield></datafield><datafield tag="035" ind1=" " ind2=" "><subfield code="a">(DE-627)NLEJ242601065</subfield></datafield><datafield tag="040" ind1=" " ind2=" "><subfield code="a">DE-627</subfield><subfield code="b">ger</subfield><subfield code="c">DE-627</subfield><subfield code="e">rakwb</subfield></datafield><datafield tag="100" ind1="1" ind2=" "><subfield code="a">Finlay, W. J. J.</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="245" ind1="1" ind2="0"><subfield code="a">Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins</subfield></datafield><datafield tag="264" ind1=" " ind2="1"><subfield code="a">Oxford, UK</subfield><subfield code="b">Blackwell Science Ltd</subfield><subfield code="c">2005</subfield></datafield><datafield tag="300" ind1=" " ind2=" "><subfield code="a">Online-Ressource</subfield></datafield><datafield tag="336" ind1=" " ind2=" "><subfield code="a">nicht spezifiziert</subfield><subfield code="b">zzz</subfield><subfield code="2">rdacontent</subfield></datafield><datafield tag="337" ind1=" " ind2=" "><subfield code="a">nicht spezifiziert</subfield><subfield code="b">z</subfield><subfield code="2">rdamedia</subfield></datafield><datafield tag="338" ind1=" " ind2=" "><subfield code="a">nicht spezifiziert</subfield><subfield code="b">zu</subfield><subfield code="2">rdacarrier</subfield></datafield><datafield tag="520" ind1=" " ind2=" "><subfield code="a">Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays.</subfield></datafield><datafield tag="533" ind1=" " ind2=" "><subfield code="d">2005</subfield><subfield code="f">Blackwell Publishing Journal Backfiles 1879-2005</subfield><subfield code="7">|2005||||||||||</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">2D SDS-PAGE</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">DeVore, N. C.</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Dobrovolskaia, E. N.</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Gam, A.</subfield><subfield code="4">oth</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Goodyear, C. S.</subfield><subfield code="4">oth</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Slater, J. E.</subfield><subfield code="4">oth</subfield></datafield><datafield tag="773" ind1="0" ind2="8"><subfield code="i">In</subfield><subfield code="t">Clinical & experimental allergy</subfield><subfield code="d">Oxford : Blackwell Science, 1989</subfield><subfield code="g">35(2005), 8, Seite 0</subfield><subfield code="h">Online-Ressource</subfield><subfield code="w">(DE-627)NLEJ243926391</subfield><subfield code="w">(DE-600)2004469-0</subfield><subfield code="x">1365-2222</subfield><subfield code="7">nnns</subfield></datafield><datafield tag="773" ind1="1" ind2="8"><subfield code="g">volume:35</subfield><subfield code="g">year:2005</subfield><subfield code="g">number:8</subfield><subfield code="g">pages:0</subfield></datafield><datafield tag="856" ind1="4" ind2="0"><subfield code="u">http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x</subfield><subfield code="q">text/html</subfield><subfield code="x">Verlag</subfield><subfield code="z">Deutschlandweit zugänglich</subfield><subfield code="3">Volltext</subfield></datafield><datafield tag="912" ind1=" " ind2=" "><subfield code="a">GBV_USEFLAG_U</subfield></datafield><datafield tag="912" ind1=" " ind2=" "><subfield code="a">ZDB-1-DJB</subfield></datafield><datafield tag="912" ind1=" " ind2=" "><subfield code="a">GBV_NL_ARTICLE</subfield></datafield><datafield tag="951" ind1=" " ind2=" "><subfield code="a">AR</subfield></datafield><datafield tag="952" ind1=" " ind2=" "><subfield code="d">35</subfield><subfield code="j">2005</subfield><subfield code="e">8</subfield><subfield code="h">0</subfield></datafield></record></collection>
|
| series2 |
Blackwell Publishing Journal Backfiles 1879-2005 |
| author |
Finlay, W. J. J. |
| spellingShingle |
Finlay, W. J. J. misc 2D SDS-PAGE Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins |
| authorStr |
Finlay, W. J. J. |
| ppnlink_with_tag_str_mv |
@@773@@(DE-627)NLEJ243926391 |
| format |
electronic Article |
| delete_txt_mv |
keep |
| author_role |
aut aut aut |
| collection |
NL |
| publishPlace |
Oxford, UK |
| remote_str |
true |
| illustrated |
Not Illustrated |
| issn |
1365-2222 |
| topic_title |
Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins 2D SDS-PAGE |
| publisher |
Blackwell Science Ltd |
| publisherStr |
Blackwell Science Ltd |
| topic |
misc 2D SDS-PAGE |
| topic_unstemmed |
misc 2D SDS-PAGE |
| topic_browse |
misc 2D SDS-PAGE |
| format_facet |
Elektronische Aufsätze Aufsätze Elektronische Ressource |
| format_main_str_mv |
Text Zeitschrift/Artikel |
| carriertype_str_mv |
zu |
| author2_variant |
a g ag c s g cs csg j e s je jes |
| hierarchy_parent_title |
Clinical & experimental allergy |
| hierarchy_parent_id |
NLEJ243926391 |
| hierarchy_top_title |
Clinical & experimental allergy |
| isfreeaccess_txt |
false |
| familylinks_str_mv |
(DE-627)NLEJ243926391 (DE-600)2004469-0 |
| title |
Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins |
| ctrlnum |
(DE-627)NLEJ242601065 |
| title_full |
Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins |
| author_sort |
Finlay, W. J. J. |
| journal |
Clinical & experimental allergy |
| journalStr |
Clinical & experimental allergy |
| isOA_bool |
false |
| recordtype |
marc |
| publishDateSort |
2005 |
| contenttype_str_mv |
zzz |
| container_start_page |
0 |
| author_browse |
Finlay, W. J. J. DeVore, N. C. Dobrovolskaia, E. N. |
| container_volume |
35 |
| physical |
Online-Ressource |
| format_se |
Elektronische Aufsätze |
| author-letter |
Finlay, W. J. J. |
| doi_str_mv |
10.1111/j.1365-2222.2005.02307.x |
| author2-role |
verfasserin |
| title_sort |
exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins |
| title_auth |
Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins |
| abstract |
Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. |
| abstractGer |
Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. |
| abstract_unstemmed |
Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays. |
| collection_details |
GBV_USEFLAG_U ZDB-1-DJB GBV_NL_ARTICLE |
| container_issue |
8 |
| title_short |
Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins |
| url |
http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x |
| remote_bool |
true |
| author2 |
DeVore, N. C. Dobrovolskaia, E. N. Gam, A. Goodyear, C. S. Slater, J. E. |
| author2Str |
DeVore, N. C. Dobrovolskaia, E. N. Gam, A. Goodyear, C. S. Slater, J. E. |
| ppnlink |
NLEJ243926391 |
| mediatype_str_mv |
z |
| isOA_txt |
false |
| hochschulschrift_bool |
false |
| author2_role |
oth oth oth |
| doi_str |
10.1111/j.1365-2222.2005.02307.x |
| up_date |
2024-07-06T02:32:32.791Z |
| _version_ |
1803795198422024192 |
| fullrecord_marcxml |
<?xml version="1.0" encoding="UTF-8"?><collection xmlns="http://www.loc.gov/MARC21/slim"><record><leader>01000caa a22002652 4500</leader><controlfield tag="001">NLEJ242601065</controlfield><controlfield tag="003">DE-627</controlfield><controlfield tag="005">20210707161450.0</controlfield><controlfield tag="007">cr uuu---uuuuu</controlfield><controlfield tag="008">120427s2005 xx |||||o 00| ||und c</controlfield><datafield tag="024" ind1="7" ind2=" "><subfield code="a">10.1111/j.1365-2222.2005.02307.x</subfield><subfield code="2">doi</subfield></datafield><datafield tag="035" ind1=" " ind2=" "><subfield code="a">(DE-627)NLEJ242601065</subfield></datafield><datafield tag="040" ind1=" " ind2=" "><subfield code="a">DE-627</subfield><subfield code="b">ger</subfield><subfield code="c">DE-627</subfield><subfield code="e">rakwb</subfield></datafield><datafield tag="100" ind1="1" ind2=" "><subfield code="a">Finlay, W. J. J.</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="245" ind1="1" ind2="0"><subfield code="a">Exploiting the avian immunoglobulin system to simplify the generation of recombinant antibodies to allergenic proteins</subfield></datafield><datafield tag="264" ind1=" " ind2="1"><subfield code="a">Oxford, UK</subfield><subfield code="b">Blackwell Science Ltd</subfield><subfield code="c">2005</subfield></datafield><datafield tag="300" ind1=" " ind2=" "><subfield code="a">Online-Ressource</subfield></datafield><datafield tag="336" ind1=" " ind2=" "><subfield code="a">nicht spezifiziert</subfield><subfield code="b">zzz</subfield><subfield code="2">rdacontent</subfield></datafield><datafield tag="337" ind1=" " ind2=" "><subfield code="a">nicht spezifiziert</subfield><subfield code="b">z</subfield><subfield code="2">rdamedia</subfield></datafield><datafield tag="338" ind1=" " ind2=" "><subfield code="a">nicht spezifiziert</subfield><subfield code="b">zu</subfield><subfield code="2">rdacarrier</subfield></datafield><datafield tag="520" ind1=" " ind2=" "><subfield code="a">Background Monoclonal antibodies are a valuable tool in the study of allergens, but the technology used in their generation can be slow and labour-intensive. Therefore, we have examined recombinant antibody development by phage-display against single allergens and protein mixtures.Objective We used the avian immunoglobulin system (generated from single VH and VL genes) to provide a rapid method for generating highly specific recombinant antibody fragments from a minimal number of animals.Methods A single-chain antibody fragment (scFv) library was generated from a single chicken immunized with model allergens. ScFvs were isolated by phage-display and their properties investigated by ELISA and Western blot.Results Mono-specific scFvs were generated against recombinant Fel d 1 and native Amb a 1. Pannings against yellow jacket venom extracts only yielded clones that reacted with multiple proteins in the venom extract. The scFvs from each panning type were effectively expressed in Escherichia coli and readily purified. Highly specific and sensitive recognition of Fel d 1 and Amb a 1 was demonstrated in ELISA, with scFvs displaying antibody-concentration-dependent absorbance curves down to picogram levels of antibody. The specificity of selected antibodies for their cognate antigen was further confirmed in Western blot analysis, with scFvs directed to either Fel d 1 or Amb a 1 showing no reactivity for the other antigens used in immunization. Anti-Amb a 1 scFvs also mapped Amb a 1-isoform location in Western blot of ragweed extracts separated by 2D SDS-PAGE. DNA sequence analysis of scFvs showed that multiple different clones had been generated against Fel d 1 and Amb a 1. Using two anti-Fel d 1 scFv for ELISA analysis of Fel d 1 content in crude cat pelt extracts, we could produce data which were highly similar (P=0.33 and 0.89 by paired t-test analysis) to those obtained using conventional assays (radial immunodiffusion).Conclusion Phage-display technology may generate multiple allergen-specific recombinant antibody fragments from a single chicken, to allergens from mammalian, plant and insect sources. The resulting antibody fragments are of demonstrable use in allergen identification and quantification, in comparison with standard immunoassays.</subfield></datafield><datafield tag="533" ind1=" " ind2=" "><subfield code="d">2005</subfield><subfield code="f">Blackwell Publishing Journal Backfiles 1879-2005</subfield><subfield code="7">|2005||||||||||</subfield></datafield><datafield tag="650" ind1=" " ind2="4"><subfield code="a">2D SDS-PAGE</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">DeVore, N. C.</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Dobrovolskaia, E. N.</subfield><subfield code="e">verfasserin</subfield><subfield code="4">aut</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Gam, A.</subfield><subfield code="4">oth</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Goodyear, C. S.</subfield><subfield code="4">oth</subfield></datafield><datafield tag="700" ind1="1" ind2=" "><subfield code="a">Slater, J. E.</subfield><subfield code="4">oth</subfield></datafield><datafield tag="773" ind1="0" ind2="8"><subfield code="i">In</subfield><subfield code="t">Clinical & experimental allergy</subfield><subfield code="d">Oxford : Blackwell Science, 1989</subfield><subfield code="g">35(2005), 8, Seite 0</subfield><subfield code="h">Online-Ressource</subfield><subfield code="w">(DE-627)NLEJ243926391</subfield><subfield code="w">(DE-600)2004469-0</subfield><subfield code="x">1365-2222</subfield><subfield code="7">nnns</subfield></datafield><datafield tag="773" ind1="1" ind2="8"><subfield code="g">volume:35</subfield><subfield code="g">year:2005</subfield><subfield code="g">number:8</subfield><subfield code="g">pages:0</subfield></datafield><datafield tag="856" ind1="4" ind2="0"><subfield code="u">http://dx.doi.org/10.1111/j.1365-2222.2005.02307.x</subfield><subfield code="q">text/html</subfield><subfield code="x">Verlag</subfield><subfield code="z">Deutschlandweit zugänglich</subfield><subfield code="3">Volltext</subfield></datafield><datafield tag="912" ind1=" " ind2=" "><subfield code="a">GBV_USEFLAG_U</subfield></datafield><datafield tag="912" ind1=" " ind2=" "><subfield code="a">ZDB-1-DJB</subfield></datafield><datafield tag="912" ind1=" " ind2=" "><subfield code="a">GBV_NL_ARTICLE</subfield></datafield><datafield tag="951" ind1=" " ind2=" "><subfield code="a">AR</subfield></datafield><datafield tag="952" ind1=" " ind2=" "><subfield code="d">35</subfield><subfield code="j">2005</subfield><subfield code="e">8</subfield><subfield code="h">0</subfield></datafield></record></collection>
|
| score |
7.399132 |