Product Description

Glycosil® is a thiol-modified hyaluronic acid and a constituent of native extracellular matrix (ECM). Glycosil is used in conjunction with Gelin-S® or ECM proteins such as laminin, collagen, or fibronectin for most 3-D cell culture and tissue-engineering applications. Tested for bacteria and endotoxins. Glycosil hyaluronic acid is a component of the HyStem®, HyStem-C and HyStem-HP hydrogel kits and can be purchased separately in individual vials.

Glycosil® (thiol-modified sodium hyaluronate) is packaged in 5 mL vials containing 50 mg. Vials are blanketed by nitrogen and under a slight vacuum.

Store Glycosil in the original vial, unopened, at -20°C for up to one year. Do not uncap the Glycosil vials since they will crosslink in the presence of oxygen. Use a syringe and needle to add DG Water and remove product from the vials through the septum.

Note: It is recommended to reconstitute each vial in its entirety.


Directions for Use

Glycosil is prepared by dissolving the lyophilized solid in the DG Water (or any sterile, degassed, deionized water). When re-constituted, it will be in 1x phosphate buffered saline (PBS), pH ~7.4. The amount of DG Water used for dissolution will depend on the vial.

Glycosil should be prepared in the following manner:

  1. Allow the Glycosil vial to come to room temperature.
  2. Under aseptic conditions, using a syringe and needle, add either 1 mL or 5 mL of DG Water to the Glycosil vial, depending on vial size (1 mL or 5 mL). 
  3. Place the vial horizontally on a rocker or shaker. It will take <40 minutes for the solids to fully dissolve. Warming to 37°C and/or gently vortexing will speed up dissolving time. Solutions will be clear and slightly viscous.
  4. Use with Extralink® or Extralink-Lite to form a hydrogel.
  5. Typically, crosslinkers are used in a 1:4 volume ratio with the polymer (i.e. 1.0 mL of Glycosil is crosslinked with 0.25 mL Extralink-Lite; or 0.5 mL Glycosil combined with 0.5 mL Gelin-S® is crosslinked with 0.25 mL Extralink). Note: The gelation time will be shorter for gels not incorporating Gelin-S. Hydrogels made using only Glycosil will not support cell attachment. Use with Gelin-S for cell attachment.

Product Q & A

Globular particles less than 75 kDa should be able to freely diffuse through a HyStem hydrogel.

When reconstituted using DG water, the pH of each HyStem component will be approximately 7.4-7.6.

One year from the date of receipt, if stored properly.

Any sterile, deionized, degassed water can be substituted for reconstitution. However, in order to ensure accurate and predictable dissolution and gelation times, our DG Water is highly recommended, as it is degassed, blanketed in argon, and has undergone validation testing with each HyStem component.

Gelin-S provides cellular attachment sites when incorporated in the hydrogel. Gelin-S is thiol-modified, denatured collagen I, derived from either bovine or porcine sources. Gelin-S is included in all HyStem-C and HyStem-HP kits.

Gelin-S has been thiol-modified in the same manner as the hyaluronan in Glycosil (or Heprasil), so that it covalently crosslinks with the Extralink in the HyStem hydrogels.

Yes. Peptides that contain a cysteine residue can be used. The cysteine residue must be present for the peptide to be covalently bonded to the hydrogel substrate.

Yes. ECM proteins, such as laminin, collagen, fibronectin, or vitronectin can be non-covalently incorporated into the hydrogel prior to crosslinking.

HyStem hydrogels and sponges differ in hydration and homogeneity. HyStem sponges are typically polymerized hydrogels that are subsequently freeze-dried. The resulting sponge is a fibrous, mesh network with pores and niches that enable cells to infiltrate and adhere. A true HyStem hydrogel is an encapsulating liquid that polymerizes around suspended cells in culture.

No. The compliance of the hydrogels is set by the amount of Extralink crosslinker added, the concentration of Glycosil (or Heprasil) and Gelin-S used, and the ratio of Glycosil (or Heprasil) to Gelin-S. Once this chemical structure of the hydrogel is fixed, it is not altered by prolonged exposure to cell culture medium.

HyStem sponges can be terminally sterilized by E-beam. HyStem hydrogels have not yet been validated for use with E-beam sterilization methods. HyStem hydrogels are not terminally sterilized by gamma irradiation.

Gelation time is affected by multiple aspects of the gel’s composition.

One way to change the gelation time of a hydrogel is to vary the amount of crosslinker used. Gels with a lower amount of Extralink crosslinker will have a longer gelation time than those with a higher amount of crosslinker. Changing the amount of crosslinker will produce slight changes in gelation time.

Gelation time can be dramatically changed by varying the Glycosil (or Heprasil) and Gelin-S concentrations. Concentrated solutions of Glycosil (or Heprasil) and Gelin-S will create a solution with a much shorter gelation time. This can easily be done by reconstituting the components in a smaller volume of DG Water. Alternatively, diluting these components in larger volumes of DG Water will dramatically increase the total time to form the hydrogel.

HyStem Hydrogels are virtually transparent and should not interfere with microscopy.

HyStem hydrogels may generate mild inflammation as part of the body’s natural healing process in response to injury. HyStem hydrogels do not trigger immune response when used in vivo. (These products are not for human use)

HyStem is degraded in vivo by matrix metalloproteinases (collagenases) and hyaluronidases.

Trypsin, Dipase, collagenase, and hyaluronidase have been used to help detach cells from the surface or from within HyStem hydrogels.

In general, the pore size for HyStem-C and HyStem-HP hydrogels is ~17 nm. 

Product Applications

Glycosil reconstituted with the recommended volume of degassed water will be a 1% solution of thiolated HA in 1X PBS salts. Upon reconstitution, Glycosil will remain liquid for several hours until crosslinker (such as Extralink or PEGSSDA) is added, at which point gelation will occur after 20-30 minutes with no temperature or pH manipulation required. Even without crosslinker, a gel will eventually form due to auto-crosslinking. Please use Glycosil within four hours of reconstitution. Diluting Glycosil with phosphate-buffered saline or cell culture media will increase gelation time or ultimately prevent gelation all together. Reconstituting Glycosil in any solvent except pure water will alter the salt balance and may impact gelation kinetics or gel properties.

Product References

References for HyStem®:

Gaetani, R., et al. (2015) Epicardial application of cardiac progenitor cells in a 3D-printed gelatin/hyaluronic acid patch preserves cardiac function after myocardial infarction. Biomaterials 61: 339-348. PMID: 17335875.

Prestwich, G.D., et al. (2007) 3-D culture in synthetic extracellular matrices: new tissue models for drug toxicology and cancer drug discovery. Adv Enzyme Regul 47: 196-207. PMID: 17335875.

Shu, X.Z., et al. (2006) Synthesis and evaluation of injectable, in situ crosslinkable synthetic extracellular matrices for tissue engineering. J Biomed Mater Res A 79: 901-912. PMID: 16941590.

Shu, X.Z., et al. (2003) Disulfide-crosslinked hyaluronan-gelatin hydrogel films: a covalent mimic of the extracellular matrix for in vitro cell growth. Biomaterials 24: 3825-3834. PMID: 12818555.

S. Cai, et al. (2005) Injectable glycosaminoglycan hydrogels for controlled release of human basic fibroblast growth factor.Biomaterials, 26, 6054-6067.

D. B. Pike, et al. (2006) Heparin-regulated release of growth factors in vitro and angiogenic response in vivo to implanted hyaluronan hydrogels containing VEGF and bFGF. Biomaterials, 27, 5242–5251.

G. D. Prestwich, et al. (2007) 3-D Culture in Synthetic Extracellular Matrices: New Tissue Models for Drug Toxicology and Cancer Drug Discovery. invited, Adv. Enz. Res., in press (2007).

X. Z. Shu, et al, (2006) Synthesis and Evaluation of Injectable, In Situ Crosslinkable Synthetic Extracellular Matrices (sECMs) for Tissue Engineering. J. Biomed Mater. Res. A, 79A(4), 901-912.

Shu, X.Z., et al. (2004) In situ crosslinkable hyaluronan hydrogels for tissue engineering. Biomaterials 25: 1339-1348. PMID: 14643608. 

Mehra, T.D., et al. (2006) Molecular stenting with a crosslinked hyaluronan derivative inhibits collagen gel contraction. J Invest Dermatol 126: 2202-2209. PMID: 16741511. 

Shu, X.Z., et al. (2004) Attachment and spreading of fibroblasts on an RGD peptide-modified injectable hyaluronan hydrogel. J Biomed Mater Res A 68: 365-375. PMID: 14704979. 

Ghosh, K., et al. (2007) Cell adaptation to a physiologically relevant ECM mimic with different viscoelastic properties. Biomaterials 28: 671-679. PMID: 17049594.

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Certificate of Origin

Safety Data Sheet

Product Disclaimer

This product is for R&D use only and is not intended for human or other uses. Please consult the Material Safety Data Sheet for information regarding hazards and safe handling practices.