CE Current Problem

Current Problems on Capillary Electrophoresis Unstable current can be caused by a cracked or damaged capillary, dirty electrodes, dirty pre-punches, or an electronics problem. First test the capillary by removing the existing capillary and installing a new one: 1. Removing the Capillary Cassette from the HP 3DCE Instrument

A. Choose the Cassette icon in the HPCE Diagram and select Change Cassette from the menu. This removes the buffer vials from the ends of the capillary. If this is not done, buffer, which may be transferred via the capillary into the electrode inserts, precipitates and may block the electrodes or damage the capillary. Open the top cover to access the cassette. Press the button in the middle of the lid front to release the latch.

B. Move the cassette guide to the front. C. Carefully pull the cassette out of the cassette guide.

2. Removing the Capillary from the Cassette

WARNING: Wear safety glasses when opening the cassette and handling the capillary to protect your eyes.

A. Open the capillary cassette by pushing the cassette lock, located in the center of the cassette.

B. Carefully remove the capillary by pushing the white capillary lock in and lifting the capillary out of the lock.

C. Uncoil the capillary from the spokes of the cassette. Put the insertion tool on a flat surface. You can also use the tool integrated into the capillary cassette.

This document is believed to be accurate and up-to-date. However, Agilent Technologies, Inc. cannot assume responsibility for the use of this material. The information contained herein is intended for use by informed individuals who can and must determine its fitness for their purpose.

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D. Put the alignment interface on the insertion tool and press down firmly. Press only on the outer ring to release the black plastic spring.

E. Hold the capillary at the colored stopper. Pull the capillary out of the alignment interface. Do not release the alignment interface while the capillary is moving. The capillary may break.

3. Installing a New Capillary in the Alignment Interface

The alignment interface must correspond to the type of capillary used. There are special alignment interfaces for straight capillaries and for extended light path capillaries. The alignment interface must also correspond to the inner diameter of the capillary. To facilitate matching of capillary and corresponding alignment interface we use the following color coding on both the sleeve of the alignment interface and the plastic stop on the capillary.

Black G1600-60150 25 m id capillaries with extended light path Green G1600-60210 50 m id straight capillaries Red G1600-60230 50 m id capillaries with extended light path Blue G1600-60310 75 m id straight capillaries Yellow G1600-60330 75 m id capillaries with extended light path

A. Make sure the color of the sleeve in the alignment interface matches the color of

the plastic stop on the capillary. B. Remove the protective plastic cap which covers the detection window.

CAUTION: Do not touch the capillary at the detection window or the capillary tips, where the polyimide is burnt off. The capillary is very fragile at that point and it is important to keep the detection window clean.

C. Put the insertion tool on a flat surface for example, your lab bench. D. Put the alignment interface on the insertion tool and press down firmly. Press

only on the outer ring to release the spring that secures the capillary in the alignment interface. You can also use the insertion tool integrated into the capillary cassette.

E. Slide the capillary through the opening in the direction of the arrow printed on the interface.

F. The capillary has a plastic stop fitted. Pull the capillary through the alignment interface until the stop is reached.

G. While still slightly pulling the capillary stop against the alignment interface, release the alignment interface from the installation tool to secure the capillary in the alignment interface.


The detection window or the extended light path of the capillary must be visible through the detection interface. If aligned correctly, you can see the slit of the interface behind the detection window. 4. Install the Capillary into the Cassette

A. Hold the capillary at the alignment interface. B. Insert the alignment interface into the alignment interface holder of the cassette.

Make sure the flat side of the alignment interface is at the outer edge of the cassette.

C. Make sure the capillary slides into the capillary guiding slit. D. Wind the capillary around the reel in the center of the capillary cassette. The

pins surrounding the reel have grooves to secure the capillary. E. Start with the inside of the upper pin to avoid the capillary being squeezed when

the cassette is closed. F. Alternate the sides of the pins when inserting the capillary.

NOTE: Make sure the windings of the capillary do not come into contact with each other. As the heat cannot be led away at contact points they get hot and may cause separation and detection problems.

G. Insert the other end of the capillary into the guiding hook at the right. H. Press the white capillary lock towards the center of the cassette and insert the

capillary into the capillary lock. Release the capillary lock to secure the capillary. I. Make sure the two capillary ends outside the cassette are both the same length

as the cassette guiding pins and are parallel to the pins. Use a sheet of paper or another soft item with a straight line to check this. Use the capillary lock to make adjustments at the inlet side.

5. Closing the Capillary Cassette

A. Connect the hinge at the side of the cassette body, which has the detection interface installed, to the one of the cassette cover.

B. Make sure the hinge is closed firmly at that side. C. Connect the other sides and latch them together. D. Make sure the cassette cover and body form one unit. E. Make sure the two capillary ends outside the cassette are both the same length

as the cassette guiding pins and are parallel to them. Use the capillary lock to make adjustments.

6. Inserting the Cassette

A. Insert the cassette into the guiding slit of the cassette guide. Make sure the ends of the capillary are not bent.


B. Slowly lower the cassette until it reaches the insulation plate. While lowering the cassette, insert the capillary ends into the capillary seats of the electrodes.

WARNING: Make sure the ends of the capillary are not bent when lowering the capillary cassette. They should slide directly into the capillary seats, otherwise they might break or be damaged. If you have problems inserting the capillary into the electrodes, check if they are blocked with salt crystals. Refer to Cleaning the Electrodes, Pre-punchers and Insulation Plate below for instructions on how to clean the electrodes.

C. Move the cassette guide back again while holding down the cassette. The

alignment interface aligns the capillary automatically to the detector. D. Close the top cover and secure it by latching it onto the body of the case. Make

sure the latch is tightened. For safety reasons no power is applied if the top cover is open. Check in upper right corner of the HPCE diagram whether the top cover is closed.

Because the cassette was changed, the HP 3D CE ChemStation automatically displays the Wavelength Calibration screen. It tests the wavelength calibration of your detector and advises you to do adjustments if necessary. Choose the Help button for more information on this feature. After you have finished Wavelength Calibration, you can close the Change Cassette dialog box by choosing Done.

E. Condition the new capillary and rerun a sample or standard to see if the current problem is resolved.

If it is not resolved, the next step will be to clean the Electrodes, Pre-punches, and Insulation Plate: 7. Cleaning the Electrodes, Pre-punches, and Insulation Plate The following materials will be needed: 12 mm hexagonal socket screwdriver (included in the startup kit). Pozidriv screwdriver (included in the startup kit). Wash bottle with isopropanol. Wash bottle with water. Compressed inert gas, oil-free. WARNING: Follow the common safety regulations for laboratories when doing this task. Wear safety spectacles and rubber gloves. Prior to maintenance do the following:


A. Select the Detector icon in the HPCE Diagram screen. B. Select Lamp Off from the menu to switch the lamp off. C. Select Maintenance from the Instrument menu to lower all lifts. D. Select Exit from the File menu to exit the HP 3D CE ChemStation. Exit both the

online and offline copy. Turn off line power to the HP 3D CE instrument. E. Disconnect the line power cord from the HP 3D CE instrument. F. Open the top cover. G. Remove the capillary cassette. H. Take off the detector cover by opening the two push turn locks. To open press

down and turn them counterclockwise.

WARNING If you were using the HP 3D CE instrument shortly before, the lamp may be very hot. Avoid touching the lamp or wear protective gloves.

8. Remove the Insulation Plate

A. Use a Pozidriv screwdriver to unscrew the two polypropylene screws which secure the insulation plate.

B. Use a Pozidriv screwdriver to remove the screw that secures the blue ground cable.

C. Use the 12 mm hexagonal socket screwdriver to unscrew the inlet electrode. D. Carefully lift the red high voltage wire. The inlet electrode will come out. E. Slowly slide the insulation plate to the right, tilt it up and take it out. Make sure

that the outlet electrode does not scrape against the foam of the tray or the lift station as you remove the plate.

F. Remove the outlet electrode from the insulation plate using the 12 mm hexagonal socket screwdriver.

G. Flush the electrodes with water. All salt crystals must be removed. H. Flush the electrodes with isopropanol. I. Use compressed air to dry the electrodes and remove ALL traces of liquid.

CAUTION: To avoid contamination, the inside of the electrodes must be completely dry before reinstallation. If there is still an accumulation of dirt in the upper funnel start the cleaning procedure again or replace the electrode.

9. Remove the Front Cover

A. Loosen the two screws that secure the front cover. It is not necessary to remove the screws completely.


B. Carefully pull the front cover away from the mainframe. Pull steadily at both sides. You need slightly more force to remove the right side due to a connector.

10. Remove the Pre-punches Inlet and outlet lifts are located behind the sample tray. The replenishment lift is located at the right side under the replenishment needle. WARNING: Do not touch or bend the replenishment needle. WARNING: The pre-punchers have a very sharp tip. Be careful when touching them.

A. Remove the plastic screw which secures the inlet pre-puncher using a Pozidriv screwdriver.

B. The pre-puncher has a hole in the side of the air tube connector which can be used to gently lever it out using a paper clip. Carefully pull the pre-puncher out of the lift.

WARNING Do not damage the funnel surface of the pre-puncher or the silicon air pressure tube connection.

C. Carefully remove the air pressure tube from the inlet pre-puncher. D. Follow the same procedure to remove the pre-puncher from the outlet lift. Check

for damaged tips before cleaning the pre-punchers. If the tips are damaged, replace the pre-punchers.

E. Flush the pre-punchers with water. Watch for salt crystals. All salt crystals must be removed. Take particular care of the air tube connector.

F. Flush the pre-punchers with isopropanol. G. Use compressed air to dry the pre-punchers and remove all traces of liquid. If

dirt is still accumulated in the pre-puncher funnel, put the prepuncher in a beaker with water and place it in an ultrasonic bath for 5 minutes followed by the isopropanol flush, and then dry with the compressed air.

CAUTION To avoid contamination the prepunchers must be completely dry before reinstallation.

11. Clean the top and bottom of the Insulation Plate

A. Moisten a tissue with water and use it to clean the insulation plate. B. Moisten a tissue with isopropanol and use it to clean the insulation plate.


C. Dry the insulation plate with tissue and compressed air. The insulation plate can also be cleaned in a dishwasher at a maximum temperature of 60 C.

CAUTION: To avoid arcing, the insulation plate must be completely dry before reinstallation.

12. Reinstall the Pre-punches

A. Carefully insert the pre-puncher labeled 8 into the outlet lift. Connect the tube to its connector (labeled 8). The fitting of the outlet pre-puncher must be tight. Otherwise problems during CEC mode may occur.

B. Carefully insert and tighten the plastic screw to secure the pre-puncher. The pre-puncher will assume its correct position as the screw is tightened.

C. Carefully connect the inlet pre-puncher with the air pressure tube (labeled 7) connected into the inlet lift. Connect the tube to its connector (labeled 7). The fitting of the inlet pre-puncher must be tight. Otherwise pressure or injection problems may result.

D. Carefully insert and tighten the plastic screw to secure the pre-puncher. The pre-puncher will assume its correct position as the screw is tightened.

13. Reinstall the Electrodes and Insulation Plate

A. Reinstall one electrode into the outlet electrode position of the insulation plate. B. Thread the electrode into place by hand first. C. Tighten the electrode with the 12 mm hexagonal socket screwdriver. Do not

overtighten. Do not bend the electrodes. D. Holding the insulation plate by the cassette holder, slide the base plate under

the detector then tilt it up into place and push it completely to the left. E. The insulation base plate should be laying flat in position. The outlet electrode

should be positioned correctly under the detector. F. Secure the base plate by reinstalling the two plastic screws. G. Reconnect the blue ground wire and fasten the screw. H. Insert the inlet electrode into the ring connector of the red high voltage wire.

Thread the inlet electrode (and the ring connector) into the base plate by hand then tighten the inlet electrode with the 12 mm hexagonal socket screwdriver. Do not overtighten.

I. Reinstall the front cover and fasten the two screws. J. Reinstall the detector cover and fasten the two push turn locks by pushing down

and turning them clockwise. K. Reinstall the capillary cassette. L. Close the top cover.



M. Connect the power cord and turn on line power to the HP 3D CE instrument. N. Start your computer and HP 3D CE ChemStation. O. Rerun sample or standard to determine if the current problem has been fixed. If

not, the problem may be electrical. Call Agilent Technologies, 1-800-227-9770, for more assistance.

Current Problems on Capillary Electrophoresis

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CE Current Problem