Pfizer researchers discuss the performance of the ExpressHT-Ultra in high-throughput DMPK studies
ExpressHT-Ultra for high speed LC/MS
Advantages of Microbore Chromatography
Low-Flow LC/MS/MS for Quantitative Bioanalysis
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introducing the cHiPLC™-nanoflex system making nanoLC/MS easier and more reproducible
Eksigent’s new cHiPLC-nanoflex, in combination with the nanoLC-Ultra system, delivers superior sensitivity, column-to-column reproducibility, and exceptional ease of operation.
The cHiPLC-nanoflex system is a “docking station” for up to three microfluidic chips. The system’s flexible design and built-in 10-port nano valve allow for easy switching between different types of experiments such as direct injection and trap-loading.
Chips containing a nanoLC column or trap column can be exchanged in seconds. A perfect dead volume free connection is made every time as the cHiPLC column or trap column is automatically aligned with a special connector chip.
All alignment is achieved through the use of highly reproducible microfabrication techniques similar to those used in the microelectronics industry. Additionally, these techniques are used to define fluidic paths and to create a microfabricated weir structure for stationary phase particle retention. Chips are temperature controlled to guarantee reproducible retention times and improved separations.
Compared with nanoLC columns constructed from fused silica tubing with end-frits made by sintering stationary phase particles, the cHiPLC columns are much more robust and far easier to handle and connect without introducing dead volumes.
Eksigent has worked on microfluidics since its start in 2000, and has incorporated chip-based products such as UV flow cells and filters in its micro-LC products. The cHiPLC-nanoflex now applies this technology to making nanoLC more reproducible and easier.
Benefits of the cHiPLC™-nanoflex system
Column design
Special care has been given to the design of both the trap-chips and analytical column-chips in order to achieve separations that are equal or better than separations obtained using packed capillaries (see Figure 1). The use of fused silica allows for cylindrical channels for packing nanoLC columns and traps. Instead of conventional frits made out of fused stationary phase particles, our cHiPLC columns use a unique weir structure to retain the stationary phase particles in the column. These weirs are more reproducible to fabricate, while their dead-volume is virtually zero (~13 pL). In addition, adsorption of sample components that can occur with frit material is not an issue with these types of structures.
Figure 1. Comparison of the separation of a peptide test mix on a nano cHiPLC column and a conventional nanoLC column. Both columns are 15 cm x 75 µm, and packed with ChromXP C18-CL 3 µm 300Å. Flowrate is 250 nl/min; gradient slope 2% Acetonitrile/min.
Patented connection system
Connections to and from each chip are made using a patented connection system that can connect up to seven channels to the outside world with a dead volume of less than 1 nl. The force used to connect the chip is pre-set, so that every time the user exchanges a chip, a leak-free connection is obtained without any required user adjustments.
Increased column-to-column reproducibility
Besides the ease of replacing a nanoLC column or trap in seconds, the use of our cHiPLC columns also increases column-to-column reproducibility. All chips are exactly identical, and our packing procedure guarantees the best possible column-to-column reproducibility in nanoLC (see Figure 2). This is of importance for applications where retention time stability over longer periods of time and over multiple columns is important. Examples are the use of retention time in combination with accurate mass in peptide/protein identification and scheduling MRMs for peptide quantitation in biomarker validation.
Figure 2. Excellent inter-column reproducibility is achieved for a peptide test mix between three 15 cm x 75 µm nano cHiPLC columns packed with ChromXP C18-CL 3 µm 300Å. Flowrate is 250 nl/min; gradient slope 2% Acetonitrile/min.
Simple operation
The cHiPLC-nanoflex allows for easy switching between direct injection, trap-loading and dual column with direct injection experiments. This is achieved through the use of a fluidic jumper chip, which routes fluid appropriately for the desired experiment. This jumper chip is as easily changed as the column and trap chips. Figure 3 shows a schematic of the set-up for a trap-loading experiment, showing the jumper, trap and column chip and the 10-port nano valve.
Figure 3. Schematic showing the set-up of the cHiPLC-nanoflex for a trap loading experiment.
Nano-spray source compatibility
In addition, the cHiPLC-nanoflex can be used in combination with all Eksigent nanoLC systems along with any mass spectrometer/nanospray source.
Figure 4. Separation of a BSA digest on a nano cHiPLC column of 15 cm x 75 µm, packed with ChromXP C18-CL 3 µm 300Å. Flowrate is 250 nl/min; gradient slope 0.5% Acetonitrile/min.
Figure 5.
Sample:
BSA tryptic digest
Column:
75 µm x 150 mm, ReproSil-Pur C18-AQ 3 µm, 120 Å
NanoLC:
Eksigent NanoLC-2D
Autosampler:
Eksigent AS1
MS:
AB SCIEX API4000 in MRM mode (70 transitions).
Data Courtesy of: Andrew James and Lorne Taylor.
Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON Canada
Dimensions
6.5” (17 cm) x 10” (25 cm) x 10.5” (26 cm) (Width x Depth x Height)
Weight
10 lbs (5 kg)
Power
100-240 VAC, 50/60 Hz, 2.5 A
Working temperature
15 to 30 ºC
Temperature control
25 to 60 ºC +/- 0.1 ºC
Maximum pressure
4000 psi
Valve
PAEK 10 port valve
1/32” connections; 100 µm bore
Port-to-port volume < 25 nl
Max. pressure 5,000 psi
Wetted parts
PEEK, PAEK, Valcon E, fused silica, tefzel
Column dimensions
75 µm ID x 15 cm
Trap column dimensions
200 µm ID x 0.5 mm
Instrument control All temperature control is achieved through the LCD interface on the nanoflex; valve control is achieved through the Eksigent software which controls the pump system. Valve control can be disabled at the nanoflex through the LCD interface.
cHiPLC™-nanoflex system ordering information
description
part number
cHiPLC nanoflex
950-00070
Nano cHiPLC column 75 µm x 15 cm ChromXP C18-CL 3 µm 120 Å
804-00001
Nano cHiPLC column 75 µm x 15 cm ChromXP C18-CL 3 µm 300 Å
804-00003
Nano cHiPLC column 75 µm x 15 cm ChromXP C18-CL 5 µm 120 Å
804-00002
Nano cHiPLC column 75 µm x 15 cm ChromXP C18-CL 5 µm 300 Å
804-00004
Nano cHiPLC Trap column 200 µm x 0.5 mm ChromXP C18-CL 3 µm 120 Å
804-00006
Nano cHiPLC Trap column 200 µm x 0.5 mm ChromXP C18-CL 3 µm 300 Å
804-00008
Nano cHiPLC Trap column 200 µm x 0.5 mm ChromXP C18-CL 5 µm 120 Å
804-00007
Nano cHiPLC Trap column 200 µm x 0.5 mm ChromXP C18-CL 5 µm 300 Å
804-00009
Direct injection jumper chip
800-00408
Trap-elute jumper chip
800-00389
Dual column jumper chip
800-00421
Chip based particle trap (weir) (between autosampler and cHiPLC nanoflex)
introducing the cHiPLC™-nanoflex system
The cHiPLC-nanoflex system is a “docking station” for up to three microfluidic chips. The system’s flexible design and built-in 10-port nano valve allow for easy switching between different types of experiments such as direct injection and trap-loading. 
