HybriWell™ Sealing System

HYBRIWELL ™ SEALING SYSTEM

HybriWell ™ Sealing System bonds securely and rapidly to a microscope slide surface to confine small reagent volumes with samples. HybriWells™ are manufactured using a clear plastic polycarbonate cover (0.010”- 0.254mm thick) and double-sided adhesives (chamber). Two access ports (1.5mm diameter) are added to each chamber to allow for the addition or removal of reactants. Ports are easily sealed using adhesive Seal Tabs (200 tabs are included), see the video. Sealed chambers are watertight and ideally suited for submerged or overnight incubations. RNase free, hydrophobic surfaces will not trap or bind probes. Press-fit tubing connectors provide affordable and easy means to connect HybriWells™ to pumps and other devices for fluidic exchange. If you are using Cy5 or ® Alexa Fluor 647 direct-labeled DNA probes please refer to our new “fluorescent friendly” chambers, see Fluorescent Friendly adhesive tab for more information. All current designs may be fabricated using our “fluorescent friendly” adhesive material.

Chamber depth and dimensions could easily be customized, a variety of adhesive options are also available for applications that employ the use of “exotic” surface chemistries. Grace Bio-Labs, has a large portfolio of adhesive options, plastics, and know-how to tailor a solution to fit your needs.

 

Fluorescent Friendly Adhesive

Standard versions of the HybriWell™ are manufactured using double-sided, clear SecureSeal™ Adhesive. The Fluorescent Friendly Adhesive (black) was developed in response to reports from few laboratories that they were experiencing signal quenching using CY5 and AF647 using SecureSeal™ Adhesive. We contacted multiple HybriWell™ customers using these fluors for hybridization and results were mixed with many of the labs having no issues whatsoever. After creating a matrix of variables, the information we know to date is;

  • The phenomenon only occurs during hybridization assays above 55C
  • Only occurs if the fluor is directly conjugated to the probe (no problem in a sandwich assay).
  • CY3 and other fluors are not affected.
  • Some labs do not have the quenching issue at all under any of the above conditions.

As far as we know, some component of the hybridization buffer is reacting with some component of the adhesive used to make standard HybriWell™ and SecureSeal™ products resulting in signal quenching. We are not able to identify which component may be the culprit as many of the customers having the problem were using pre-made hybe buffers, the manufacturers of which will not share the formulation with us.  We feel that it is appropriate to alert customers to the potential risk of this quenching phenomenon if they are using SecureSeal™ adhesive under any of the conditions we’ve described. We identified the Fluorescent Friendly adhesive as a material which does not quench signal under any conditions and it was tested by the labs which reported the problem and all reported favorable results.

Technical Resources

Instructions

Instructions for Use HybriWell

Applications

  • In situ hybridization
  • Protein and DNA Microarrays
  • Immunocytochemistry
  • DNA Amplification
  • Neurophysiology
  • Reaction kinetics
  • Rapid microfluidic prototyping
  • Nucleic Acid Programmable Protein Arrays
  • Fluorescence Resonance Energy Transfer (FRET)
  • Single molecule fluorescence microscopy

References

Vang S, Seitz K, Krysan PJ. "A simple microfluidic device for live cell imaging of Arabidopsis cotyledons, leaves, and seedlings." Biotechniques. 2018 Jun;64(6):255-261. 

 2018 Jun;64(6):255-261. doi: 10.2144/btn-2018-0044.

Siebman, Coralie, Orlin D Velev, and Vera I Slaveykova. "Two-Dimensional Algal Collection and Assembly by Combining AC-Dielectrophoresis with Fluorescence Detection for Contaminant-Induced Oxidative Stress Sensing." Biosensors 5, no. 2 (2015): 319-36.

Jeenduang, Nutjaree, Sureerut Porntadavity, Markus Von Nickisch-Rosenegk, Frank F Bier, and Chamras Promptmas. "Two-dye Based Arrayed Primer Extension for Simultaneous Multigene Detection in Lipid Metabolism.Clinica Chimica Acta; International Journal of Clinical Chemistry 442 (2015): 36-43.
 

Helmer, Dorothea, Ina Rink, James A. R. Dalton, Kevin Brahm, Marina Jöst, Tobias M. Nargang, Witali Blum, et al. “Rational Design of a Peptide Capture Agent for CXCL8 Based on a Model of the CXCL8:CXCR1 Complex.RSC Adv. 5, no. 33 (2015): 25657–68. doi:10.1039/C4RA13749C.

To see more references for this product click here.

Videos

HybriWell Sealing System Tubing Connectors for Loading Reagents and Perfusion Applications

FAQ

Q: What is the best way to change solutions within a HybriWell™ chamber if I want to leave it affixed to the slide?

A: The simplest method to remove solutions from a HybriWell™ is to un-cap both filling ports, tip the slide vertically and allow the solution to drain.  For faster exchange or if the reagent is viscous, un-cap both filling ports and place the chambered slide into a 50ML conical tube.  Give it a quick low-speed spin (1000 rpm) in the centrifuge and all of the solution will be collected in the bottom of the tube.  This, by the way, is a great method to recover and re-use some primary antibodies or collect pre-hybe buffer to which you can add the probe, mix and load back to the chamber. 

Q: What is the difference between HybriWell™ and SecureSeal™?

A: Fundamentally, the difference between a HybriWell™ and a SecureSeal™ Chamber is the surface to volume ratio created by the differences in chamber depth.   To achieve the best active mixing,  select a chamber size in which the fluid moves freely during rotation.  Rotational mixing can be further facilitated by leaving a small “mixing bubble” within the chamber.  SecureSeals™ use greater fluid volume per surface area compared to  HybriWells™, however, to conserve reagents. HybriWells™ may be the better choice.  Both products are recommended for DNA chip slide chemistries, considering the reagents used, incubation time and temperature and surface modification of the glass, one type of chamber may bond more aggressively than the other, as they use different adhesives assemblies.

Q: I am trying to do in situ hybridization and we have 2 mouse tissue sections per slide.  Are there individual HybriWells™ where I can put one over each section on the slide?

A: Yes, we have small single-well HybriWells™, and two of them will fit on a microscope slide to create two different chambers. See our HybriWell™ 8-pack assortment (item #615101) which contains HybriWells™ of various sizes that can be adapted to your application.  Each of the HybriWells™ in the 8-pack Assortment is also sold individually in packs.