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GPC4-Nanobody - stem cell differentiation booster

https://www.idylle-labs.com/web/image/product.template/678/image_1920?unique=a0e24f2

Single-domain antagonizing antibody blocking Glypican-4 protein activity and promoting stem cell entry into differentiated lineages

390.00 € 390.0 EUR 390.00 €

390.00 €

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A technology developed by Rosanna Dono (Institut du Biologie du Développement de Marseille IBDM, Aix-Marseille Université, France)

GPC4-Nanobody is a single-domain antagonizing antibody (sdAb) that recognizes and binds a conformational epitope of the native human Glypican-4 (hGPC4) with high affinity and specificity. When added to the culture medium of live cells, GPC4-Nanobody blocks hGPC4 biological functions. 

GPC4-Nanobody can be used as a differentiation booster: by maintaining reduced hGPC4 levels throughout differentiation, GPC4-Nanobody addition in the differentiation medium leads to more efficient differentiated lineage entry in response to triggering factors. 


A differentiation booster

GPC4-Nanobody arises from the groundbreaking discovery that constant reduction in hGPC4 levels in differentiating hiPSCs makes them more responsive to differentiation signals (cf Publications section). Continuous exposure of hiPSC to GPC4-Nanobody allows them to acquire a unique biologial state characterized by:

  • Maintenance of pluripotency in undifferentiated conditions (absence of differentiation factors)
  • Enhanced differentiation yields upon differentiation exposure 
  • Reduced tumorigenicity in xenografts


Applications

GPC4 nanobody can be added directly into the culture medium to:

-> Increase differentiation yields of human stem cells for mechanistic studies or subsequent in vivo transplantation 

So far, GPC4-Nanobody has been shown to increase differentiation yield of hiPSC into endoderm progenitors and dopaminergic neurons.

-> Detect hGPC4 

GPC4-Nanobody can be used to detect and quantify hGPC4 levels (native or recombinant) with high specificity in immunostaining, flow cytometry or immunoprecipitation assays performed unfer non-denaturing conditions. 

This nanobody binds to hGPC4 only in its native conformation: it cannot be used for hGPC4 detection in Western Blots or other assays performed under denaturing conditions. 

-> Block hGPC4 

GPC4-Nanobody binds hGPC4 with high affinity and blocks its biological functions: beyond hGPC4 detection, it can also be used to specifically inhibit its activity temporally or permanently in live cells. A simple way to unravel hGPC4 specific roles in various biological contexts without inducing any genetic modifications in mechanistic studies or therapeutic/diagnostic tool development!     

Kit contents

Vial contents: Lyophilized GPC4-Nanobody 

Available formats: 2 mg, 6 mg or 12 mg 

Storage instructions: the lyophilized nanobody can be stored at room temperature for up to one year. Once reconstituted, aliquot and freeze for long-term storage. 


Working concentration: 50 - 500 nM 


GPC4-Nanobody increases endoderm differentiation efficiency in human induced pluripotent stem cells (hiPSCs)









Left: Immunofluorescence analysis of SOX17 endodermal marker (red) and DAPI (blue) positive

cells in hiPSC cultured for 2 days with endodermal differentiation factors alone (Control) or added with GPC4-Nanobody at increasing concentrations. 

Right: Quantification of the percentage of SOX17+ progenitors over DAPI+ (n = 4 biological replicates).

Credits: Rosanna Dono (Institut du Biologie du Développement de Marseille IBDM, Aix-Marseille Université, France)


Downregulation of GPC4 in hiPSCs promotes efficient generation of ventral midbrain dopamine neuron progenitors in vitro





A. Immunofluorescence analysis of LMX1A (red), FOXA2 (green), OTX2 (purple), and DAPI (blue) positive

cells in CTRLsh, GPC4sh5-c10, and GPC4sh4-c8 cultures at the indicated time points of VMDA differentiation. LMX1A+, FOXA2+, OTX2+ cells,

which correspond to bona fide VMDA progenitors, are highlighted by dashed areas with a star. B. Quantification of the percentage of LMX1A+,

FOXA2+, OTX2+ VMDA progenitors over DAPI+ (VMDA progenitors at d9: CTRLsh: 8.1 ± 1.8%; GPC4sh5-c10: 26.5 ± 1.6%; GPC4sh4-c8:

42.3 ± 3.1%; at d12: CTRLsh: 13.3 ± 1.2%; GPC4sh5-c10: 27.1 ± 3.1%; GPC4sh4-c8: 31.4 ± 4.6%; at d15: CRTLsh: 31.4 ± 3.1%; GPC4sh5-c10:

62.7 ± 7.6%; GPC4sh4-c8: 62.7 ± 9.0%). Data are presented as mean ± SEM (n = 2-3 biological replicates). One-way ANOVA: *P < .05, **P < .01,

***P < .001.

Source publication: Corti S. et al, 2020

Downregulation of GPC4 in hiPSCs enhances differentiation potential into definitive endoderm


A. Immunofluorescence analysis of SOX17 (red, DE) onWT, CTRLsh, GPC4sh5-c10 and GPC4sh2-c3 029 hiPSCs differentiated for 3 days into DE, n = 3. Scale bar: 100 μm. B. Percentages of SOX17 positive cells in WT, CTRLsh, GPC4sh5-c10 and GPC4sh2-c3 029 hiPSCs were quantified from staining shown in A. Box plots represent the median with min and max values, n = 3. Statistical analysis: two-way ANOVA, followed by Dunnett’s multiple comparison test. P-values: (***) <0.001, (**) <0.01, (*) <0.05, ns not significant. 

Source publication: Legier et al, 2023


Remi Bonjean, Brigitte Kerfelec, Patrick Chames, Rosanna Dono, Functional targeting of Glypican-4 by a conformation-specific single-domain antibody, bioRxiv 2025.05.10.653258; doi: https://doi.org/10.1101/2025.05.10.653258

Legier, T., Rattier, D., Llewellyn, J. et al. Epithelial disruption drives mesendoderm differentiation in human pluripotent stem cells by enabling TGF-β protein sensing. Nat Commun 14, 349 (2023). https://doi.org/10.1038/s41467-023-35965-8

Corti S, Bonjean R, Legier T, Rattier D, Melon C, Salin P, Toso EA, Kyba M, Kerkerian-Le Goff L, Maina F, Dono R. Enhanced differentiation of human induced pluripotent stem cells toward the midbrain dopaminergic neuron lineage through GLYPICAN-4 downregulation. Stem Cells Transl Med. 2021 May;10(5):725-742. doi: 10.1002/sctm.20-0177. Epub 2021 Feb 2. PMID: 33528918; PMCID: PMC8046045.
Publication highlighted on the IBDM Institute's website: https://www.ibdm.univ-amu.fr/a-new-target-to-control-the-differentiation-and-tumorigenic-potentials-of-human-induced-pluripotent-stem-cells/

GPC4-Nanobody is a patented technology (International Publication Number: WO 2022/079270 A1)

Everspark technology has been intially developed by Karine Monier, Arnaud Favier and Christophe Place and and published in Scientific Reports:
Provost, A., Rousset, C., Bourdon, L. et al. Innovative particle standards and long-lived imaging for 2D and 3D dSTORM. Sci Rep 9, 17967 (2019). https://doi.org/10.1038/s41598-019-53528-0
 

Publications:

Nanoscale engagement and clusterization of Programmed death ligand 1 (PD-L1) in the membrane lipid rafts of Non-Small Cell Lung Cancer cells 
Martina Ruglioni, Simone Civita, Tiziano Salvadori, Sofia Cristiani, Vittoria Carnicelli, Serena Barachini,  Iacopo Petrini, Irene Nepita, Marco Castello, Alberto Diaspro, Paolo Bianchini, Barbara Storti, Ranieri Bizzarri,  Stefano Fogli and Romano Danesi   bioRxiv 2022.08.09.503318; doi: https://doi.org/10.1101/2022.08.09.503318

HIV-1 diverts actin debranching mechanisms for particle assembly and release in CD4 T lymphocytes
Rayane Dibsy, Erwan Bremaud, Johnson Mak, Cyril Favard, Delphine Muriaux
BioRxiv, December 16, 2022. doi. 10.1101/2022.12.15.520580

Fluorescent Polymer-AS1411-Aptamer Probe for dSTORM Super-Resolution Imaging of Endogenous Nucleolin
Fabre L, Rousset C, Monier K, Da Cruz-Boisson F, Bouvet P, Charreyre MT, Delair T, Fleury E, Favier A. Biomacromolecules. 2022 May 12. doi: 10.1021/acs.biomac.1c01706. PMID: 35549176

Comparative analysis of ChAdOx1 nCoV-19 and Ad26.COV2.S SARS-CoV-2 vector vaccines.
Michalik S, Siegerist F, Palankar R, Franzke K, Schindler M, Reder A, Seifert U, Cammann C, Wesche J, Steil L, Hentschker C, Gesell-Salazar M, Reisinger E, Beer M, Endlich N, Greinacher A, Völker U. Haematologica. 2022 Apr 1;107(4):947-957. doi: 10.3324/haematol.2021.280154. PMID: 35045692

Metabolic biorthogonal labeling and dSTORM imaging of peptidoglycan synthesis in Streptococcus pneumoniae 
Jennyfer Trouve, Oleksandr Glushonkov and Cecile Morlot
Star Protocols, December 13, 2021. doi: 10.1016/j.xpro.2021.101006

Insights in ChAdOx1 nCoV-19 vaccine-induced immune thrombotic thrombocytopenia.
Greinacher A, Selleng K, Palankar R, Wesche J, Handtke S, Wolff M, Aurich K, Lalk M, Methling K, Völker U, Hentschker C, Michalik S, Steil L, Reder A, Schönborn L, Beer M, Franzke K, Büttner A, Fehse B, Stavrou EX, Rangaswamy C, Mailer RK, Englert H, Frye M, Thiele T, Kochanek S, Krutzke L, Siegerist F, Endlich N, Warkentin TE, Renné T. 
Blood. 2021 Dec 2;138(22):2256-2268. doi: 10.1182/blood.2021013231. PMID: 34587242

Superresolution Microscopy of Drosophila Indirect Flight Muscle Sarcomeres.
Szikora S, Novák T, Gajdos T, Erdélyi M, Mihály J. Bio Protoc. 2020 Jun 20;10(12):e3654. doi: 10.21769/BioProtoc.3654. eCollection 2020 Jun 20. PMID: 33659324