About: A novel strategy for efficient production of anti-V3 human scFvs against HIV-1 clade C

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About: A novel strategy for efficient production of anti-V3 human scFvs against HIV-1 clade C Goto Sponge NotDistinct Permalink

An Entity of Type : schema:ScholarlyArticle, within Data Space : wasabi.inria.fr associated with source document(s)

Attributes	Values
type	Academic Article research paper schema:ScholarlyArticle
isDefinedBy	Covid-on-the-Web dataset
has title	A novel strategy for efficient production of anti-V3 human scFvs against HIV-1 clade C
Creator	Kumar, Rajesh Wig, Naveet Andrabi, Raiees Luthra, Kalpana Sinha, Subrata Dutta, Durgashree Khan, Lubina Sankhyan, Anurag Tiwari, Ashutosh Prakash, Somi
Source	PMC
abstract	BACKGROUND: Production of human monoclonal antibodies that exhibit broadly neutralizing activity is needed for preventing HIV-1 infection, however only a few such antibodies have been generated till date. Isolation of antibodies by the hybridoma technology is a cumbersome process with fewer yields. Further, the loss of unstable or slowly growing clones which may have unique binding specificities often occurs during cloning and propagation and the strongly positive clones are often lost. This has been avoided by the process described in this paper, wherein, by combining the strategy of EBV transformation and recombinant DNA technology, we constructed human single chain variable fragments (scFvs) against the third variable region (V3) of the clade C HIV-1 envelope. RESULTS: An antigen specific phage library of 7000 clones was constructed from the enriched V3- positive antibody secreting EBV transformed cells. By ligation of the digested scFv DNA into phagemid vector and bio panning against the HIV-1 consensus C and B V3 peptides followed by random selection of 40 clones, we identified 15 clones that showed V3 reactivity in phage ELISA. DNA fingerprinting analysis and sequencing showed that 13 out of the 15 clones were distinct. Expression of the positive clones was tested by SDS-PAGE and Western blot. All the 13 anti-V3 scFvs showed cross-reactivity against both the clade C and B V3 peptides and did not show any reactivity against other unrelated peptides in ELISA. Preliminary neutralization assays indicated varying degrees of neutralization of clade C and B viruses. EBV transformation, followed by antigen selection of lines to identify specific binders, enabled the selection of phage from un-cloned lines for scFv generation, thus avoiding the problems of hybridoma technology. Moreover, as the clones were pretested for antigen binding, a comparatively small library sufficed for the selection of a considerable number of unique antigen binding phage. After selection, the phage clones were propagated in a clonal manner. CONCLUSIONS: This strategy can be efficiently used and is cost effective for the generation of diverse recombinant antibodies. This is the first study to generate anti-V3 scFvs against HIV-1 Clade C.
has issue date	2012-11-15(xsd:dateTime)
bibo:doi	10.1186/1472-6750-12-87
bibo:pmid	23153214
has license	cc-by
sha1sum (hex)	ef64fa65c77a221698f7cf13acdcf801ed2d554b
schema:url	https://doi.org/10.1186/1472-6750-12-87
resource representing a document's title	A novel strategy for efficient production of anti-V3 human scFvs against HIV-1 clade C
has PubMed Central identifier	PMC3536577
has PubMed identifier	23153214
schema:publication	BMC Biotechnol
resource representing a document's body	covid:ef64fa65c77a221698f7cf13acdcf801ed2d554b#body_text
is schema:about of	named entity 'LOSS OF' named entity 'PRODUCTION' named entity 'RECOMBINANT DNA TECHNOLOGY' named entity 'MONOCLONAL ANTIBODIES' named entity 'GROWING' named entity 'CLONES' named entity 'CHAIN' named entity 'CLONING' named entity 'SLOWLY' named entity 'MAY HAVE' named entity 'ACTIVITY' named entity 'COMBINING' named entity 'HIV-1' named entity 'LOST' named entity 'BACKGROUND' named entity 'STRONGLY POSITIVE' named entity 'GENERATED' named entity 'ANTIBODIES' named entity 'EBV TRANSFORMATION ' named entity 'CLADE' named entity 'STRATEGY' named entity 'SINGLE' named entity 'PAPER' named entity 'IS A' named entity 'HYBRIDOMA' named entity 'EFFICIENT' covid:arg/ef64fa65c77a221698f7cf13acdcf801ed2d554b named entity 'CLADE' named entity 'STRATEGY' named entity 'HUMAN' named entity 'NOVEL' named entity 'PRODUCTION' named entity 'ENVELOPE' named entity 'CONSTRUCTED' named entity 'EXHIBIT' named entity 'VARIABLE' named entity 'BINDING' named entity 'PROPAGATION' named entity 'PROCESS' named entity 'TILL' named entity 'HIV-1' named entity 'DATE' named entity 'UNIQUE' named entity 'OCCURS DURING' named entity 'ISOLATION' named entity 'TECHNOLOGY' named entity 'HAVE' named entity 'SPECIFICITIES' named entity 'HIV-1 INFECTION' named entity 'VARIABLE REGION' named entity 'PREVENTING' named entity 'DESCRIBED' named entity 'NEEDED' named entity 'HUMAN' named entity 'UNSTABLE' named entity 'FRAGMENTS' named entity 'antibodies' named entity 'variable region' named entity 'cloning' named entity 'scFvs' named entity 'HIV-1' named entity 'antibodies' named entity 'HIV-1' named entity 'scFvs' named entity 'antibodies' named entity 'antibody' named entity 'viruses' named entity 'phage display technology' named entity 'NaCl' named entity 'pH 9' named entity 'ligation' named entity 'scFvs'

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