1801 YAPLSRL(48) KLWVIPQ(33) 2 Phage display (competitive panning) 3 PMID:15574836 70 kDa heat shock cognate protein 1 Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) ELISA The binding ability of the high-frequency peptides was confirmed by ELISA. Absorbances at 405 nm were measured (data not shown). The heptapeptide KLWVIPQ displayed about 60% of the binding ability to Vr330 as compared with YAPLSRL, which had the highest binding ability. The relative binding ability of Vr330 to any other low-frequency heptapeptide was also <5%. The recombinant proteins were designated Vr330 for polypeptides containing the C-terminal 30 kDa domain of VrHsc70-3. The recombinant proteins were purified on a metal-affinity column and then cleaved with thrombin to remove the His tag. The Vr330 recombinant proteins was used to select VrHsc70-binding heptapeptides using phage display. The bound phage was eluted with a 1 ml solution of the free target protein. Among the 116 different clones of Vr330, 41.4% were peptides with a sequence of YAPLSRL and 28.4% had the peptide sequence of KLWVIPQ. The heptapeptide KLWVIPQ displayed about 60% of the binding ability to Vr330 as compared with YAPLSRL, which had the highest binding ability confirmed by ELISA. 1802 EPGHDAVP E-X(2)-H 1 Phage display (competitive panning) 4 PMID:16398491 Cobalt ion, Co(2+) Not determined. Not determined. Not determined. Type 8 phage display library (X8) The selection of type 8 phage with affinity toward Co2+ was performed by incubating the type 8 phage library (~e10 pfu) with Co2+ immobilized on 200 μL of Chelating Sepharose Fast Flow gel in TBS buffer (pH 7.5) with 0.15% tween-20. The bound phage were washed 10 times with the incubation buffer and then eluted with 50 mM histidine. 1803 WHWRLPS(10)[2.290 ± 0.094] HKPLQIW(5)[0.363 ± 0.088] QAHTVGK(4)[0.834 ± 0.451] YVHRLPP(2)[0.555 ± 0.078] HYTRHSA(1)[0.373 ± 0.114] 5 Phage display (common panning) 3 PMID:16413648 Microcystin-LR, MC-LR Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) ELISA Affinities for MC-LR of the five different selected clones were measured by ELISA. Absorbances at 410 nm were reproduced from the bar graph and expressed as the means of three determinations ± sample standard deviations. L7 represented original linear 7 peptides library and its absorbance was 0.256 ± 0.266. Clone L7-3 (displaying WHWRLPS) expressed the highest affinity for MC-LR as compared to the original library and their negative controls. The affinity of selected phage to MC-LR were tested through an immunoprecipitation assay. The results showed that all selected phage combined with MC-LR except the YVHRLPP-bearing phage. 1804 GRIQRWWVMFLL(1) IISRAGVVRGNE(1) ADYLSRWGSIRN(1) ETYVFDNHFHAP(1) TVSEVASTSNLD(1) SRVHRAVLNGVS(1) RPPGVVRRYALG(1) VRSWEEQARVTT(1) RAFIASRRIKRP(1) VSQRVGFITSAP(1) AGAGGGNVPVCS(1) RESTLKGTSRAV(1) AGLRLKKAAIHR(1) SSLLRAVPEPTG(1) EWDITTECTVTF(1) 15 Bacterial display (common panning) 10 PMID:16599553 Petri dish (polystyrene, PS) Not determined. Not determined. Not determined. FliTrx bacterial display library (X12) ELISA The mutant-type GSTs fused with PS-tags no. 16, 17, 19, and 23 (SRVHRAVLNGVS, RPPGVVRRYALG, RAFIASRRIKRP and AGLRLKKAAIHR) were adsorbed to the polystyrene latex beads and the percentage was 80% or higher, whereas those for the wild-type GST and the GST-PS1 were only 50%. RAFIASRRIKRP and AGLRLKKAAIHR showed a strong affinity to PS latex beads with relatively high residual activities. 1805 KGLRGWREMISL(5) LDPGAMRTIVRP(3) QLVEGVRHRIWN(2) 3 Bacterial display (common panning) 5 DOI:10.1016/j.molcatb.2003.12.019 96-well immunoassay plate (polystyrene, PS) Not determined. Not determined. Not determined. FliTrx bacterial display library (X12) The selected peptides were commonly rich in hydrophobic amino acid residues and had two or three basic amino acid residues. Adsorption and desorption experiments for one of the selected peptide (KGLRGWREMISL) showed that it was well and irreversibly adsorbed onto PS latex particles. The most frequent sequence was LGP*AAR*FGDE, where the asterisks represent stop codons. This sequence is not included because of the insertions of stop codons. 1806 LPPWQRQ(9)[0.476 ± 0.014] HPERATL(8)[0.492 ± 0.031] KPRMPPR(4)[0.500 ± 0.028] HDHRYPK(2)[0.494 ± 0.007] HPRWHTP(1)[0.455 ± 0.013] QLKTGLA(1)[0.423 ± 0.033] GKPMPPM(1)[0.337 ± 0.025] THLPWQT(1)[0.382 ± 0.015] HPWWRPS(1)[0.440 ± 0.012] HALGPSS(1)[0.472 ± 0.019] HAIYPRH(1)[0.477 ± 0.004] HKPDANR(1)[0.483 ± 0.018] AITRSPA(1)[0.357 ± 0.014] FPGHSGP(1)[0.400 ± 0.013] NKNYIQH(1)[0.350 ± 0.021] 15 Phage display (common panning) 3 PMID:17154206 Isotactic poly(methyl methacrylate), st-PMMA Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) ELISA A triplicate ELISA was performed to measure the affinities of all selected clones to conditioned st-PMMA. Absorbances at 405 nm were determined, reproduced from the graph and shown as means ± standard deviations. The absorbance for negative control was 0.120 ± 0.012. st-PMMA films, approximately 40-nm thick, were spincoated on glass slides and immersed in Tris-buffered saline (TBS, Tris=tris(hydroxymethyl)aminomethane; pH 7.4) for 15 h (conditioning). The relative amount of phages bound to the PMMA films is quantified by ELISA. Significantly, for many clones, greater amounts bind to target films than to reference films. The amounts bound to reference films were almost the same within experimental error. 1807 HPERATL(10)[0.492 ± 0.031] LPPWQRQ(3)[0.476 ± 0.014] HPVHPHR(1)[0.507 ± 0.024] 3 Phage display (common panning) 4 PMID:17154206 Isotactic poly(methyl methacrylate), st-PMMA Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) ELISA A triplicate ELISA was performed to measure the affinities of all selected clones to conditioned st-PMMA. Absorbances at 405 nm were determined, reproduced from the graph and shown as means ± standard deviations. The absorbance for negative control was 0.120 ± 0.012. st-PMMA films, approximately 40-nm thick, were spincoated on glass slides and immersed in Tris-buffered saline (TBS, Tris=tris(hydroxymethyl)aminomethane; pH 7.4) for 15 h (conditioning). The relative amount of phages bound to the PMMA films is quantified by ELISA. Significantly, for many clones, greater amounts bind to target films than to reference films. The amounts bound to reference films were almost the same within experimental error. 1808 HPERATL(14)[0.492 ± 0.031] LPPWQRQ(1)[0.476 ± 0.014] 2 Phage display (common panning) 5 PMID:17154206 Isotactic poly(methyl methacrylate), st-PMMA Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) ELISA A triplicate ELISA was performed to measure the affinities of all selected clones to conditioned st-PMMA. Absorbances at 405 nm were determined, reproduced from the graph and shown as means ± standard deviations. The absorbances for negative control was 0.120 ± 0.012. st-PMMA films, approximately 40-nm thick, were spincoated on glass slides and immersed in Tris-buffered saline (TBS, Tris=tris(hydroxymethyl)aminomethane; pH 7.4) for 15 h (conditioning). The relative amount of phages bound to the PMMA films is quantified by ELISA. Significantly, for many clones, greater amounts bind to target films than to reference films. The amounts bound to reference films were almost the same within experimental error. 1809 HPERATL(30)[0.492 ± 0.031] LPPWQRQ(10)[0.476 ± 0.014] HPRWHTP(1)[0.455 ± 0.013] HPRLGLA(1)[0.413 ± 0.033] 4 Phage display (common panning) 8 PMID:17154206 Isotactic poly(methyl methacrylate), st-PMMA Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) ELISA A triplicate ELISA was performed to measure the affinities of all selected clones to conditioned st-PMMA. Absorbances at 405 nm were determined, reproduced from the graph and shown as means ± standard deviations. The absorbances for negative control was 0.120 ± 0.012. st-PMMA films, approximately 40-nm thick, were spincoated on glass slides and immersed in Tris-buffered saline (TBS, Tris=tris(hydroxymethyl)aminomethane; pH 7.4) for 15 h (conditioning). The relative amount of phages bound to the PMMA films is quantified by ELISA. Significantly, for many clones, greater amounts bind to target films than to reference films. The amounts bound to reference films were almost the same within experimental error. 1810 MPLQRLFDGASP(2)[1.308 ± 0.033/1.534 ± 0.047] AYPPSLYDGYNP(1)[1.134 ± 0.027] STHAKLSDGSNP(1)[1.221 ± 0.024] DDRLTSRDGSNP(1)[1.284 ± 0.060] YQLRPNAESLRF(1)[1.161 ± 0.043] GQPLIAHSTKLL(1)[1.534 ± 0.044] YALTPVPSSIRT(1)[1.527 ± 0.043] LFDGSNP 7 Phage display (common panning) 3 PMID:18485511 Anti-E2 protein monoclonal antibody HQ06 E2 protein Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA The selected peptides for target binding was detected by phage ELISA. Three independent assays were carried out. Absorbances at 490 nm were determined, reproduced from the graph and shown as means of three independent assays ± standard deviations. Phage ELISA results showed that the selected eight phage clones all showed specific reactivity with HQ06 (OD490 nm > 1.10), but not with anti-porcine IFN-γ mAb (OD490 nm < 0.15, data not shown). The present study describes the identification of a linear B-cell epitope at the N-terminus of the E2 protein by screening a phage-displayed random 12-peptide library with the neutralizing monoclonal antibody HQ06 directed against the E2 protein. HQ06 was found to bind to the phages displaying a consensus motif LFDGSNP, which is highly homologous to 772LFDGTNP778 of the CSFV polyprotein, locating on the border between antigenic domains B/C and A of the E2 protein. 1811 APWHLSSQYSRT NLAPIKVSLTSL SNQIPSSARAFI TALATSSTYDPH 4 Phage display (common panning) 5 PMID:18565751 Sulfated Lewis X Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) Panning was carried out in a 96-well Streptavidin High Binding Capacity coated microplate or 20 μL of streptavidin agarose bead slurry in a disposable column. 1812 FAAPMRTVQKID RNRRSIQRPMIS 2 Phage display (common panning) 5 PMID:18565751 Sulfated Lewis A Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) Panning was carried out in a 96-well Streptavidin High Binding Capacity coated microplate or 20 μL of streptavidin agarose bead slurry in a disposable column. The synthesized peptide FAAPMRTVQKID displayed a consistently high binding affinity and specificity against the cognate HSO3-LeA. The dimeric FAAPMRTVQKID peptide has a low micromolar affinity against the sulfated Lewis A, which is even more specific than an antibody. 1813 WDANGKT(1) ASSLNIA(4) LAPQKLP(1) VSAAPYP(1) AAANVWH(1) AYPGFAL(1) TATITTK(1) 7 Phage display (in vivo) 5 PMID:10204780 Mouse skeletal muscles Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) Initially, three rounds of in vitro selection were accomplished on differentiated murine C2C12 myotubes. Phage selected after three in vitro rounds on C2C12 mouse myotubes were injected intravenously into mice to screen for muscle binding phage in vivo. Skeletal muscles were harvested, and the phage rescued from the tissues were amplified and rescreened in a second round. After five rounds of enrichment, phage were plated, 10 random phage clones were picked from the plate. Sequencing the corresponding coding region in the viral DNA from muscle-localizing phage revealed one dominant peptide, ASSLNIA, representing 40% of the clones. Phage carrying this peptide showed a 9- to 20-fold (depending on control tissue) increase in muscle selectivity compared with phage with no insert. When the injected individual phage clone was localized by immunohistochemistry, it was found within focal areas of the membrane of myofibers. Thus, the peptide identified represents a ligand that is capable of accessing skeletal and cardiac muscle from the lumen of blood vessels and could therefore readily be exploited for targeted delivery to muscle cells. 1814 TPAWLDPPT(12) PTAQRWRFR(1) 2 Phage display (common panning) 3 PMID:18502181 Anti-human DAF monoclonal antibody DG3 Complement decay-accelerating factor Not determined. Not determined. pVIII-9aa phage display library (X9) ELISA A total of 100 clones were randomly chosen after the third round of screening and were subjected to ELISA. Thirteen clones were found to react strongly with DG3. The specificity of the two different peptides to immobilized DAF was confirmed by competitive ELISA. The results showed that phage 2 harboring peptide sequences TPAWLDPPT, but not phage 1 harboring peptide sequences PTAQRWRFR , inhibited the binding of DG3 to immobilized DAF in a concentration-dependent manner, demonstrating the specificity of phage 2 to DG3. Virions of the phage library were incubated for 1 h at room temperature with 10 μg of affinitypurified anti-DAF DG3 antibodies. To capture the phage-antibody complexes, 0.5μg/mL biotinylated goat anti-mouse IgG was incubated for 30 min with the antibody-phage complexes. The Petri dish coated with 3μg streptavidin/mL was washed four times and the phage-antibody complexes were added and incubated for 90 min at 37 °C. A total of 100 clones were randomly chosen after the third round of screening and were subjected to ELISA. Thirteen clones were found to react strongly with DG3. All these phage clones were sequenced. These 13 clones comprised only two different sequences. One sample was an independent clone (PTAQRWRFR, frequency 1/13, phage 1), and the other 12 shared the same sequence (TPAWLDPPT, frequency 12/13, phage 2). Two isolated sequences did not show obvious homology. The amino acid sequence similarity between the selected clones and DAF was investigated. Although no absolute consensus sequences were observed between phage 2 and the SCR3 of DAF, phage 2 showed sequence similarity with amino acids 210-218 of DAF SCR3, To determine whether the peptide TPAWLDPPT mimics the structure of the DG3-binding epitope of DAF, competitive binding of DG3 and the peptide TPAWLDPPTwith immobilized DAF was investigated by ELISA. The binding of DG3 to DAF was inhibited by adding the peptide TPAWLDPPT in a concentration-dependent manner, while the control peptide (DAF SCR3 homologous peptide, SVQWSDPPT) had no effect on the binding of DG3 to DAF. These data support that the peptide TPAWLDPPT mimics the DG3-binding epitope of DAF, and that DG3 recognizes a nonlinear constrained epitope. 1815 RSNTRMTARQHRSANHKSTQRARS(8) RSVFLPSILGWRSRLDDQGVAARS(2) RSTRNKHTTARRSVAPGIGEPSRS(3) RSIMHVRLRARRSARHMKDADPRS(1) RSPIIIRSRINRSHGRTKATPARS(1) RSRGLRNILMLRSYDSRSMRPHRS(2) RSTRRGTHNKDRS(4) RSTVPKRHPKDRS(1) RSIAKKTHNKQRS(1) RSYDSRSMRPHRS(1) RSTASRHTEPHRS(1) 11 Bacterial display (common panning) PMID:10618196 Zinc oxide, ZnO Not determined. Not determined. Not determined. X9 E. coli bacterial display library ZnO and bacteria expressing the random peptide library in FimH were mixed and allowed to adhere to each other at room temperature with gentle agitation. Centrifugation was then performed, and the ZnO and any adhering bacteria were recovered and inoculated into Luria-Bertani medium containing the appropriate antibiotics. After overnight incubation, exponentially growing cultures were established, and the enrichment procedure was repeated. The sequences selected exhibited various degrees of affinity and specificity towards ZnO. Competitive binding experiments revealed that the sequences recognized only the oxide form of Zn. Interestingly, one of the inserts, RSNTRMTARQHRSANHKSTQRARS, exhibited significant homology to a specific sequence in a putative zinc-containing helicase, which suggests that searches such as this one may aid in identifying binding motifs in nature. 1816 CNRPFIPTC 1 Phage display (subtractive panning, competitive panning) 4 PMID:12565725 Prothrombin Hirudin variant-2 1A2C,1A46,1A5G,1A61,1B5G,1K21,1K22,1NRS,1VZQ,1W7G,1WAY,2C8W,2C8X,2C8Y,2C8Z,2C90,2C93,2R2M,2ZFQ,2ZFR,2ZG0,2ZHE,2ZHF,2ZHW,3BIU,3BIV,3F68,3HTC,4HTC,1A4W,1DOJ,1TMB,1TMU,3HAT,3P17,3QTO,3QTV,3QWC,3QX5,3RLW,3RLY,3RM0,3RM2,3RML,3RMM,3RMN,3RMO,3SHA,3SHC,3SI3,3SI4,3SV2,3T5F,3TU7,3U98,3U9A,3UWJ,4BAO,4E7R,4LOY,4N3L,4NZE, Not determined. Ph.D.-C7C phage display library (CX7C) Phages were added to half of the blocked streptavidin magnetic particles. The phage supernatant was then transferred to the thrombin-bound magnetic particles coated with streptavidin and incubated overnight at room temperature. In the first round, phages bound to thrombin were eluted with 50 μl of 0.1 M glycine, pH 2, for 15 min at room temperature and immediately neutralised with 125 μl of 1 M Tris-HCl, pH 8. The titre of the eluted phages was estimated and half of the eluted fraction was used for amplification. Three additional rounds of panning were performed. During this last three rounds, the thrombin-bound phages were eluted with 8 mg r-hirudin (Hoechst-Marion-Rousell, AGS Frankfurt, Germany). This peptide, thrombin-inhibiting peptide (TIP), is a full competitive inhibitor of thrombin with an inhibition constant (Ki) of 0.4974 mM. It lengthened the thrombin time and inhibited thrombin-induced platelet activation and the platelet release reaction, both in a dose-dependent manner. It also reduced platelet adhesion onto a human microvascular endothelial matrix in the parallel plate flow chamber under both arterial and venous shear conditions. 1817 TSFAEYWNLLSP(6)[3.4][3.3] LTFEHYWAQLTS(6)[NT][20] DDWFQRVWSPLM(1)[NT][NT] RYEFLDYWSQLH(1)[NT][NT] VPRSAPTLWLGT(1)[NT][NT] F-X(3)-W-X(2)-L 5 Phage display (common panning) 4 PMID:19255450 p53-binding domains of MDM2 Cellular tumor antigen p53 1YCR,4HFZ, 3EQS,3LNZ,3G03,3JZR,3JZS, Ph.D.-12 phage display library (X12) Surface plasmon resonance (SPR),Isothermal titration calorimetry (ITC) A surface plasmon resonance (SPR)-based competition assay was performed with (15–29)p53 as the competitor. The Kd value (nM) of synthesized peptides was shown. Besides, the direct interaction between p53-binding domains of MDM2 (referred to thereafter as (syn)MDM2) and the peptide (TSFAEYWNLLSP) were quantified at 25°C, using a MicroCal VP-ITC microcalorimeter. The binding affinity (Kd, nM) was shown. NT denotes not tested. The p53-binding domains of MDM2 (referred to thereafter as (syn)MDM2) and their site-specifically biotinylated forms were chemically synthesized using native chemical ligation. In fact, biotin-(syn)MDM2 was used as bait against the phage display library. Several MDM2 structures have been determined in complexes with PMI (PDB ID code 3EQS and 3LNZ) and pDI (PDB ID code 3G03, 3JZR and 3JZS). The basic procedures for library screening are as follows: (i) incubate input phage (10 μL) with 400 μL of 10 nM biotin (syn)MDM2 for 60 min before adding phage-target solution to 50 μL of streptavidin-agarose resin (Pierce) for affinity capturing; (ii) wash unbound phage; (iii) elute bound phage with 1 mM (15–29) p53; (iv) amplify the eluate and collect phage for the next round of panning; (v) repeat steps 1– 4; (vi) sequence selected binding clones according to the procedures recommended by the manufacturer. Two consensus sequences emerged for both MDM2 and MDMX: LTFEHYWAQLTS, also termed pDI, and PMI–TSFAEYWNLLSP. The 3 most critical residues involved in p53-MDM2/MDMX recognition, i.e., Phe-19, Trp-23 and Leu-26 (p53 numbering), were all present in the phage-selected consensus sequences. The most potent inhibitor (TSFAEYWNLLSP), termed PMI, bound to MDM2 and MDMX at low nanomolar affinities—approximately 2 orders of magnitude stronger than the wild-type p53 peptide of the same length (ETFSDLWKLLPE). Comparative structural analysis identified an extensive, tightened intramolecular H-bonding network in bound PMI that contributed to its conformational stability, thus enhanced binding to the 2 oncogenic proteins. Importantly, the C-terminal residue Pro of PMI induced formation of a hydrophobic cleft in MDMX previously unseen in the structures of p53-bound MDM2 or MDMX. 1818 LTFEHYWAQLTS(7)[77][NT] TSFAEYWNLLSP(5)[4.2][8.9] DDWFQRVWSPLM(1)[NT][NT] NTFREYWNQLPT(1)[NT][NT] DDWFQRVVSPLM(1)[NT][NT] F-X(3)-W-X(2)-L 5 Phage display (common panning) 4 PMID:19255450 p53-binding domains of MDMX Cellular tumor antigen p53 3DAB, 3EQY,3FDO,3JZO,3JZP,3JZQ, Ph.D.-12 phage display library (X12) Surface plasmon resonance (SPR),Isothermal titration calorimetry (ITC) A surface plasmon resonance (SPR)-based competition assay was performed with (15–29)p53 as the competitor. The Kd value (nM) of synthesized peptides was shown. Besides, the direct interaction between p53-binding domains of MDM2 (referred to thereafter as (syn)MDM2) and the peptide (TSFAEYWNLLSP) were quantified at 25°C, using a MicroCal VP-ITC microcalorimeter. The binding affinity (Kd, nM) was shown. NT denotes not tested. The p53-binding domains of MDMX (referred to thereafter as (syn)MDMX) and their site-specifically biotinylated forms were chemically synthesized using native chemical ligation. In fact, biotin-(syn)MDMX was used as bait against the phage display library. The basic procedures for library screening are as follows: (i) incubate input phage (10 μL) with 400 μL of 10 nM biotin (syn)MDMX for 60 min before adding phage-target solution to 50 μL of streptavidin-agarose resin (Pierce) for affinity capturing; (ii) wash unbound phage; (iii) elute bound phage with 1 mM (15–29) p53; (iv) amplify the eluate and collect phage for the next round of panning; (v) repeat steps 1– 4; (vi) sequence selected binding clones according to the procedures recommended by the manufacturer. Two consensus sequences emerged for both MDM2 and MDMX: LTFEHYWAQLTS, also termed pDI, and PMI–TSFAEYWNLLSP. The 3 most critical residues involved in p53-MDM2/MDMX recognition, i.e., Phe-19, Trp-23 and Leu-26 (p53 numbering), were all present in the phage-selected consensus sequences. The most potent inhibitor (TSFAEYWNLLSP), termed PMI, bound to MDM2 and MDMX at low nanomolar affinities—approximately 2 orders of magnitude stronger than the wild-type p53 peptide of the same length (ETFSDLWKLLPE). Comparative structural analysis identified an extensive, tightened intramolecular H-bonding network in bound PMI that contributed to its conformational stability, thus enhanced binding to the 2 oncogenic proteins. Importantly, the C-terminal residue Pro of PMI induced formation of a hydrophobic cleft in MDMX previously unseen in the structures of p53-bound MDM2 or MDMX. Several MDMX structures have been determined in complexes with PMI (PDB ID code 3EQY) and pDI (PDB ID code 3FDO, 3JZO, 3JZP and 3JZQ). 1819 VDWVGWGASW(5) WVEWRRGWLS(1) SRTWVLWIRY(4) WGYVFHGWNL(3) MWGMHWAGEW(2) CCWGYGGSLV(1) 6 Phage display (common panning) 3 PMID:9048419 Polystyrene/polyurethane magnetic beads Not determined. Not determined. Not determined. fUSE5-based X10 phage display library Twenty randomly chosen clones from the third round of selection were analyzed further. Only clones binding significantly better to the M-280 beads than control phages from the unselected library or a wild-type phage without insert are shown. 1820 PWNAWGRTWV(5) VYRVIWFVGR(1) GGWAMWGRVW(2) FHAFHAWSAA(1) RPVAGDHPGS(1) GSVWWQAHVDR(1) WRLGFARWIG(3) WMRWGRSWGR(1) GSWKEWGGAW(1) LNWRGGWNWF(1) 10 Phage display (subtractive panning) 3 PMID:9048419 Polystyrene/polyurethane magnetic beads Not determined. Not determined. Not determined. fUSE5-based X10 phage display library A 15-mer synthetic peptide ODIN (GREPRVATVTRILRQ) was covalently attached to tosyl-activated M-280 beads by its N-terminal amino group. For counter selection, amplified phage from the first round of selection were incubated with beads coated no peptides. The supernatants were used as input to the second round. The initial rational was to apply the phage display technique to select peptides attached to magnetic beads. Individual phage clones, however, did not bind to ODIN-peptide adsorbed to microplate wells, where they bound strongly to uncoated as well as peptide-coated beads. Thus, the selection process had apparently resulted in enrichment of phages with affinity for the bead material and not for the peptide, despite deliberate attempts to eliminate plastic-binding clones from the phage pool prior to the second round of panning by counter-selection against beads without attached peptides. Twenty randomly chosen clones from the third round of selection were analyzed further. Only clones binding significantly better to the M-280 beads than control phages from the unselected library or a wild-type phage without insert are shown. 1821 SAWVRWGRVW(18) FLPYARWFSR(1) WSAWVGDWAL(6) VIYYFLVMFM(1) SRGFWLVWEQ(1) 5 Phage display (subtractive panning) 3 PMID:9048419 Polystyrene/polyurethane magnetic beads Not determined. Not determined. Not determined. fUSE5-based X10 phage display library A 15-mer synthetic peptide ODIN (GREPRVATVTRILRQ) was covalently attached to tosyl-activated M-280 beads by its N-terminal amino group. For counter selection, amplified phage from the first round of selection were incubated with beads coated no peptides. The supernatants were used as input to the second round. The initial rational was to apply the phage display technique to select peptides attached to magnetic beads. Individual phage clones, however, did not bind to ODIN-peptide adsorbed to microplate wells, where they bound strongly to uncoated as well as peptide-coated beads. Thus, the selection process had apparently resulted in enrichment of phages with affinity for the bead material and not for the peptide, despite deliberate attempts to eliminate plastic-binding clones from the phage pool prior to the second round of panning by counter-selection against beads without attached peptides. Twenty randomly chosen clones from the third round of selection were analyzed further. Only clones binding significantly better to the M-280 beads than control phages from the unselected library or a wild-type phage without insert are shown. 1822 VCFVMFSMCR(1) GVIHWRAWGGDW(1) LWGGLYGIWM(1) TWTLWFASMS(1) LSAWARWGSS(1) 5 Phage display (subtractive panning) 3 PMID:9048419 Polystyrene/polyurethane magnetic beads Not determined. Not determined. Not determined. fUSE5-based X10 phage display library A 15-mer synthetic peptide ODIN (GREPRVATVTRILRQ) was covalently attached to tosyl-activated M-280 beads by its N-terminal amino group. For counter selection, amplified phage from the first round of selection were incubated with beads coated no peptides. The supernatants were used as input to the second round. The initial rational was to apply the phage display technique to select peptides attached to magnetic beads. Individual phage clones, however, did not bind to ODIN-peptide adsorbed to microplate wells, where they bound strongly to uncoated as well as peptide-coated beads. Thus, the selection process had apparently resulted in enrichment of phages with affinity for the bead material and not for the peptide, despite deliberate attempts to eliminate plastic-binding clones from the phage pool prior to the second round of panning by counter-selection against beads without attached peptides. Twenty randomly chosen clones from the third round of selection were analyzed further. Only clones binding significantly better to the M-280 beads than control phages from the unselected library or a wild-type phage without insert are shown. 1823 PTWRSM(10) TRMRPG(1) VLLSVA(1) INQVRF(1) WCSRLF(1) PCHCSF(1) RGYFFK(1) 7 Phage display (common panning) 3 PMID:7682645 Monoclonal antibody 3-4B Not determined. Not determined. Not determined. f3-6mer phage display library (X6) ELISA The difference between A405 and A495 is taken as the signal. The strongest signals amount to ~0.6, while background is ~0.02. Only relatively strong ligands give signals in this assay. Phages displaying a ligand for the biotinylated antibodies are captured on the dish by binding of the biotin moiety to immobilized streptavidin. 1824 ATWAVL(1) AVMTSS(1) SWFLQW(1) REWISH(1) YTALLI(1) CYLCSV(1) LFSSGK(1) VWHLLH(1) VPWWVP(1) LSRILF(1) PGHSPW(1) WNLRSS(1) IALMDY(1) 13 Phage display (common panning) 3 PMID:7682645 Insulin Not determined. Not determined. Not determined. f3-6mer phage display library (X6) ELISA The difference between A405 and A495 is taken as the signal. The strongest signals amount to ~0.6, while background is ~0.02. Only relatively strong ligands give signals in this assay. Phages displaying a ligand for the biotinylated insulin are captured on the dish by binding of the biotin moiety to immobilized streptavidin. 1825 RAWSYV(1) KGRYQQ(1) EHGRPQ(1) GCSDVL(1) NLLSMT(1) CLGEHD(1) RFYGGS(1) ILPLRI(1) LSRSYF(1) NLYLVH(1) AWFRRL(1) LRGKLS(1) VDVGRS(1) 13 Phage display (common panning) 3 PMID:7682645 Interleukin-2 receptor subunit alpha Interleukin-2 (IL-2) 1Z92,2B5I,2ERJ, Not determined. f3-6mer phage display library (X6) ELISA The difference between A405 and A495 is taken as the signal. The strongest signals amount to ~0.6, while background is ~0.02. Only relatively strong ligands give signals in this assay.