Frequently Asked Questions about CelerityCE™ Bonded Capillaries
- What is the advantage of using CelerityCE™ capillaries with my proteins with buffers that are below pH 4?
- If there is a positive charge on the capillary wall at pH 4 and below, isn’t there going to be a reverse direction EOF?
- How do I separate neutral compounds on CelerityCE™ bonded capillaries?
- I have tried bonded capillaries from other suppliers and they seemed to suffer from lack of capacity. How is CelerityCE™ different?
- I have read that a limitation to this type of CE is the relatively long axial distances that molecules have to migrate to interact with the bonded moiety. How does CelerityCE™ capillary overcome this limitation?
- If I use proteins with CelerityCE™ capillaries, won’t the un-bound silanol sites on the capillary walls interact with them causing poor separations?
- What is the best way to regain flow in the capillary if I allowed it to dry out overnight, accidentally?
- Would an air bubble in the CelerityCE™ capillary be an indication of a problem?
- What would be the symptoms of a cracked CelerityCE capillary?
- What is best way to know if I have a crack in my capillary?
- What is the best way to determine if I have blockage in my CelerityCE™ Capillary?
- Can I destroy the CelerityCE™ capillary by applying too much pressure to it?
- Does my CE unit have enough pressure to force solvents and buffers through the column?
- What is the best way to condition or get the CelerityCE™ capillary ready for separation?
- What other steps should I take to ensure optimal and easy electropherogram/chromatograms with the CelerityCE capillaries?
- What procedure do you recommend for degassing solvents and buffers for use with CelerityCE™ Capillaries?
- My Agilent CE system requires a 47.5cm long capillary. You provide 70cm capillaries. That means we will have to waste more then 25% of the capillary. Why do you have so much excess?
1. What is the advantage of using CelerityCE™ capillaries with my proteins with buffers that are below pH 4? [top]
CelerityCE™ bonded capillaries have many advantages for protein separations below pH 4. One important advantage
is that if your protein’s pI is above 4, then working below this pH will make your proteins positively charged.
Below pH 4, the unique surface chemistry of CelerityCE™ makes the wall of capillary positively charged. This
means that wall adsorption is minimized or eliminated. Also, another important advantage of CelerityCE™
capillaries is that there will be both electrophoretic and chromatographic effects.
2. If there is a positive charge on the capillary wall at pH 4 and below, isn’t there going to be a reverse
direction EOF? [top]
Yes! The reverse direction EOF, even with normal polarity, helps in the separation process.
3. How do I separate neutral compounds on CelerityCE™ bonded capillaries? [top]
CelerityCE™ combines features of both chromatography and electrophoresis and therefore two separation
mechanisms: solute/bonded phase interactions (as measured by capacity factor k’) and electrophoretic mobility
(µep). For neutral molecules, only k’ interactions result in differences in migration rates among different
molecules and electrophoretic mobility does not have impact on the separations.
4. I have tried bonded capillaries from other suppliers and they seemed to suffer from lack of capacity. How is
CelerityCE™ different? [top]
CelerityCE™ capillaries are chemically etched before bonding thus increasing the surface area by one order of
magnitude. This allows more stationary phase to be bonded to the capillary walls and thereby increases the overall
capacity of the capillary. Other capillaries from other suppliers do not use the etched surface and suffered from
low capacity.
5. I have read that a limitation to this type of CE is the relatively long axial distances that molecules have to
migrate to interact with the bonded moiety. How does CelerityCE™ capillary overcome this limitation? [top]
The dissolution and re-deposition of silica material during our proprietary etching process creates radial extensions
or protrusions from the wall that decreases the distance a solute must travel in order to interact with the
stationary phase. This is especially important for large molecules such as proteins where diffusion coefficients are
relatively low.
6. If I use proteins with CelerityCE™ capillaries, won’t the un-bound silanol sites on the capillary walls
interact with them causing poor separations? [top]
Generally speaking, no! Some of the reagents used in the etching process during the manufacturing of the
CelerityCE™ capillaries become part of the surface of the wall. This new surface composition greatly reduces
the adsorptive properties of the capillary wall. Also, the patented bonding chemistry employed in CelerityCE™
manufacturing process replaces residual silanols with hydride groups below the bonded phase. Both of these aspects
makes CelerityCE™ an ideal candidate for protein separations as the wall interactions are virtually elminated.
7. What is the best way to regain flow in the capillary if I allowed it to dry out overnight, accidentally? [top]
The best way to regain flow is to remove 1 or 2mm of each end of the capillary. Then you should force methanol
through the capillary using a syringe. Click here to view a convenient hand pump for this
purpose.
8. Would an air bubble in the CelerityCE™ capillary be an indication of a problem? [top]
Yes but not with the capillary. If you develop an air bubble and you have taken care to de-gas your mobile phase, an
air bubble will not develop in a CelerityCE capillary under normal use when your instrument if working properly.
It is possible that you may have trapped some air while installing the capillary. You should “flush” the capillary
with a hand held syringe to push the air out of the capillary so that you can maintain current. Use a
small enough syringe with a piece of tubing that you can insert the capillary into. Apply
pressure and inspect under a microscope or hand held inspection-scope as the air bubble
“marches” out of the capillary.
9. What would be the symptoms of a cracked CelerityCE capillary? [top]
A crack in the CelerityCE capillary could cause your current to be very erratic and unstable. Depending on where the
crack in the capillary is, you may be able to cut the capillary and eliminate the crack.
10. What is best way to know if I have a crack in my capillary? [top]
The best way to know is to look at the entire capillary under either a stereo microscope or using a hand held
magnifier offered in our catalog. Click here to view a catalog description of the
inspection-scope.
11. What is the best way to determine if I have blockage in my CelerityCE™ Capillary? [top]
The first thing that I would do is to inspect the capillary under a stereo microscope or by using the
Inspection Scope listed in our Online Catalog. You can see through the column into the open
tubular part of the column under normal conditions. If there is a blockage, you should be able to see it.
12. Can I destroy the CelerityCE™ capillary by applying too much pressure to it? [top]
No, the "coating" is chemically bonded to the wall with a patented process that makes it very, very strong. The
ligand used should be able to withstand the same pressure and other conditions as an HPLC ligand does.
13. Does my CE unit have enough pressure to force solvents and buffers through the column? [top]
Yes, unlike the pre-packed CEC columns such as the Milli-phase™ CEC Columns, most Commercially available CE
units have sufficient capabilities of forcing solvents and mobile phases through an unblocked Celerity CE capillary.
14. What is the best way to condition or get the CelerityCE™ capillary ready for separation? [top]
The recommended procedure for preparing the MicroSolv CelerityCE™ bonded capillaries is to force the
buffer/solvent you plan to use through the capillary with a syringe. You should do this for 3-5 minutes. Then
install the capillary into your CE instrument, run with buffer/solvent and check for a current. Within a few minutes
you should see a stable baseline. No other conditioning steps are needed. Click here
to view full procedure.
15. What other steps should I take to ensure optimal and easy electropherogram/chromatograms with the CelerityCE capillaries? [top]
One of the most important steps that you can take to ensure the best results is to make sure that your
buffer/solvent used in the CelerityCE™ capillaries is thoroughly degassed.
16. What procedure do you recommend for degassing solvents and buffers for use with CelerityCE™ Capillaries? [top]
We usually recommend that to achieve optimal results with the MicroSolv CelerityCE™ capillaries, you sonicate
the buffers/solvents followed by a helium or argon purge for about 15 minutes.
17. My Agilent CE system requires a 47.5cm long capillary. You provide 70cm capillaries. That means we will have to waste more then 25% of the capillary. Why do you have so much excess? [top]
Although the Agilent system requires a 47.5cm capillary, other systems require as much as 65cm. You can take advantage of
this extra capillary in the following way: make your window in the full capillary from one end. If you break the window,
cut the part with the window off and start again. Do not cut your capillary to length then make your window.
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