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[[File:ScFv-rotation.gif|thumb|Rotating scFv fragment with highlighted complementarity determining regions (CDRs)]]
[[File:Single chain variable fragment.svg|thumb|The two possible structures of a single-chain variable fragment, with the antigen binding sites including the [[N-terminus|N-termini]] on the left and the [[C-terminus|C-termini]] on the right. The linker peptides are shown as arrows.]]
A '''single-chain variable fragment''' ('''scFv''') is not actually a [[Antibody fragment|fragment]] of an antibody, but instead is a [[fusion protein]] of the variable regions of the [[Immunoglobulin heavy chain|heavy]] (V<sub>H</sub>) and [[Immunoglobulin light chain|light chains]] (V<sub>L</sub>) of [[immunoglobulins]], connected with a short linker [[peptide]] of ten to about 25 [[amino acid]]s.<ref>{{cite journal | last1 = Huston | first1 = J. S. | last2 = Levinson | first2 = D. | last3 = Mudgett-Hunter | first3 = M. | last4 = Tai | first4 = M. S. | last5 = Novotný | first5 = J. | last6 = Margolies | first6 = M. N. | last7 = Crea | first7 = R. | year = 1988 | title = Protein engineering of antibody binding sites: recovery of specific activity in an anti-digoxin single-chain Fv analogue produced in Escherichia coli | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 85 | issue = 16| pages = 5879–5883 | doi=10.1073/pnas.85.16.5879| pmid = 3045807 | pmc = 281868 | bibcode = 1988PNAS...85.5879H | doi-access = free }}</ref> The linker is usually rich in [[glycine]] for flexibility, as well as [[serine]] or [[threonine]] for solubility, and can either connect the [[N-terminus]] of the V<sub>H</sub> with the [[C-terminus]] of the V<sub>L</sub>, or ''vice versa''.<ref name="Schirrmann">{{cite
This protein retains the specificity of the original immunoglobulin, despite removal of the constant regions and the introduction of the linker.<ref name="Peterson" /> The image to the right shows how this modification usually leaves the specificity unaltered.
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==Bivalent and trivalent scFvs==
[[File:Polyvalent single-chain variable fragments.svg|thumb|300px|Structure of divalent (top) and trivalent (bottom) scFvs, tandem (left) and di-/trimerisation format (right)]]
''Divalent'' (or ''bivalent'') single-chain variable fragments (di-scFvs, bi-scFvs) can be engineered by linking two scFvs. This can be done by producing a single peptide chain with two V<sub>H</sub> and two V<sub>L</sub> regions, yielding ''tandem scFvs''.<ref>{{cite journal|title=Development of tumor targeting anti-MUC-1 multimer: effects of di-scFv unpaired cysteine ___location on PEGylation and tumor binding|first5=SJ|last5=Denardo|first4=GL|last4=Denardo|first3=XB|last3=Shi|first2=A|last2=Natarajan |last1=Xiong|first1=Cheng-Yi|journal=Protein Engineering Design and Selection|year=2006|volume=19|issue=8|pmid=16760193|pages=359–367|doi=10.1093/protein/gzl020|doi-access=free}}</ref><ref>{{cite journal|title=A revival of bispecific antibodies|first1=Peter|last1=Kufer|first2=Ralf|last2=Lutterbüse|first3=Patrick A.|last3=Baeuerle|journal=Trends in Biotechnology|volume=22|issue=5|pages=238–244|year=2004|url=http://www.micromet.de/fileadmin/template/main/pdf/publications_147881aaf61df52304237f0ee7f0cf2a.pdf|doi=10.1016/j.tibtech.2004.03.006|pmid=15109810}}</ref> Another possibility is the creation of scFvs with linker peptides that are too short for the two variable regions to fold together (about five amino acids), forcing scFvs to dimerize. This type is known as ''diabodies''.<ref>{{cite journal|pmid=8341653|title="Diabodies": small bivalent and bispecific antibody fragments|last1=Hollinger|first1=Philipp|volume=90|issue=14|pmc=46948|date=July 1993|pages=6444–8|last2=Prospero|first2=T|last3=Winter|first3=G|journal=Proceedings of the National Academy of Sciences of the United States of America|doi=10.1073/pnas.90.14.6444|bibcode=1993PNAS...90.6444H|doi-access=free}}</ref> Diabodies have been shown to have [[dissociation constant]]s up to 40-fold lower than corresponding scFvs, meaning that they have a much higher [[affinity (pharmacology)|affinity]] to their target. Consequently, diabody drugs could be dosed much lower than other therapeutic antibodies and are capable of highly specific targeting of tumors in vivo.<ref name="Adams">{{cite journal|pmid=9652755|year=1998|last1=Adams|first1=GP|last2=Schier|first2=R|last3=McCall|first3=AM|last4=Crawford|first4=RS|last5=Wolf|first5=EJ|last6=Weiner|first6=LM|last7=Marks|first7=JD|title=Prolonged in vivo tumour retention of a human diabody targeting the extracellular ___domain of human HER2/neu|volume=77|issue=9|pages=1405–12|pmc=2150193|journal=British Journal of Cancer|doi=10.1038/bjc.1998.233}}</ref> Still shorter linkers (one or two amino acids) lead to the formation of trimers, so-called ''triabodies'' or ''tribodies''. ''Tetrabodies'' have also been produced. They exhibit an even higher affinity to their targets than diabodies.<ref>{{cite journal|title=Di-, tri- and tetrameric single chain Fv antibody fragments against human CD19: effect of valency on cell binding|first4=M|last4=Little|first3=G|last3=Moldenhauer|journal=FEBS Letters|first2=SM|volume=453|last2=Kipriyanov|issue=1|pages=164–168|first1=F.|last1=Le Gall|pmid=10403395|doi=10.1016/S0014-5793(99)00713-9|year=1999|s2cid=20213440|doi-access=free}}</ref>
All of these formats can be composed from variable fragments with specificity for two different antigens, in which case they are types of [[bispecific antibodies]].<ref>{{cite journal|pmid=11388794|year=2001|last1=Dincq|first1=S|last2=Bosman|first2=F|last3=Buyse|first3=MA|last4=Degrieck|first4=R|last5=Celis|first5=L|last6=De Boer|first6=M|last7=Van Doorsselaere|first7=V|last8=Sablon|first8=E|title=Expression and purification of monospecific and bispecific recombinant antibody fragments derived from antibodies that block the CD80/CD86-CD28 costimulatory pathway|volume=22|issue=1|pages=11–24|doi=10.1006/prep.2001.1417|journal=Protein Expression and Purification}}</ref><ref>{{cite
==Examples==
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