Waters XBridge Protein BEH, C4, 300A, 3.5 µm Columns - Manual (Page 10)

10
[ CARE AND USE MANUAL ]
XBridge Protein BEH C
4
, 300Å, 3.5, 5, 10 �m
use a Keystone, Inc. SLIPFREE ® connector to always ensure the
correct fit. The finger tight SLIPFREE connectors automatically
adjust to fit all compression screw type fittings without the use of
tools (Figure 7).
Figure 7: Single and Double SLIPFREE Connectors
SLIPFREE Connector Features:
§ Tubing pushed into endfitting, thereby guaranteeing a
void-free connection
§ Connector(s) come(s) installed on tubing
§ Various tubing i.d.s and lengths available
§ Fingertight to 10,000 psi – never needs wrenches
§ Readjusts to all column endfittings
§ Compatible with all commercially available endfittings
§ Unique design separates tube-holding function from
sealing function
Table 3: Waters Part Numbers for SLIPFREE Connectors
SLIPFREE Type Tubing Internal Diameter
Tubing
Length 0.005” 0.010” 0.020”
Single 6 cm PSL 618000 PSL 618006 PSL 618012
Single 10 cm PSL 618002 PSL 618008 PSL 618014
Single 20 cm PSL 618004 PSL 618010 PSL 618016
Double 6 cm PSL 618001 PSL 618007 PSL 618013
Double 10 cm PSL 618003 PSL 618009 PSL 618015
Double 20 cm PSL 618005 PSL 618001 PSL 618017
Minimizing Band Spreading
Figure 8 shows the influence of tubing internal diameter on
system band spreading and peak shape. As can be seen, the
larger tubing diameter causes excessive peak broadening and
lower sensitivity.
Diluted/Distorted Sample Band
0.005 inches
0.020 inches
0.040 inches
Figure 8: Effect of Connecting Tubing on System
b. Measuring System Variance and System Volume
This test should be performed on an HPLC system with a UV detector.
1. Disconnect column from system and replace with a zero dead
volume union.
2. Set flow rate to 1 mL/min.
3. Dilute a test mix in mobile phase to give a detector sensitivity
of 0.5–1.0 AUFS (system start up test mix can be used which
contains uracil, ethyl, and propyl parabens; Waters Part
Number WAT034544).
4. Inject 2 to 5 μL of this solution.
5. Measure the peak width at 4.4% of peak height
(5-sigma method):
5-sigma Bandspreading (μL) = Peak Width (min) x Flow
Rate (mL/min) x (1000 μL/1 mL)
System Variance (μL 2 ) = (5-sigma bandspreading) 2 / 25
System Volume
4.4% h
Figure 9: Determination of System Bandspreading Volume Using 5-Sigma Method