Waters XBridge Protein BEH, C4, 300A, 3.5 µm Columns - Manual (Page 9)

9
[ CARE AND USE MANUAL ]
XBridge Protein BEH C
4
, 300Å, 3.5, 5, 10 �m
1. Correct connection of 1/16-inch outer diameter stainless
steel tubing leading to and from the column is essential for
high-quality chromatographic results.
2. When using standard stainless steel compression screw
fittings, it is important to ensure proper fit of the 1/16-inch
outer diameter stainless steel tubing. When tightening or
loosening the compression screw, place a 5/16-inch wrench on
the compression screw and a 3/8-inch wrench on the hex head
of the column endfitting.
3. If a leak occurs between the stainless steel compression screw
fitting and the column endfitting, a new compression screw
fitting, tubing and ferrule must be assembled.
4. An arrow on the column identification label indicates correct
direction of solvent flow.
Correct connection of 1/16–inch outer diameter stainless steel
tubing leading to and from the column is essential for high-quality
chromatographic results. To obtain a void-free connection, the
tubing must touch the bottom of the column endfitting. It is
important to realize that extra column peak broadening due to
voids can destroy an otherwise successful separation. The choice
of appropriate column connectors and system tubing is discussed
in detail below.
0.130 inche 0.090 inches
Figure 3: Waters and Parker Ferrule Types
Due to the absence of an industry standard, various column
manufacturers have employed different types of chromatographic
column connectors. The chromatographic separation can be
negatively affected if the style of the column endfittings does not
match the existing tubing ferrule settings. This section explains
the differences between Waters style and Parker style ferrules and
endfittings (Figure 3). Each endfitting style varies in the required
length of the tubing protruding from the ferrule. The XBridge Column
is equipped with Waters style endfittings that require a 0.130-inch
ferrule depth. If a non-Waters style column is presently being used, it
is critical that ferrule depth be reset for optimal performance prior to
installing an XBridge Protein BEH C
4
, 300Å Column.
In a proper tubing/column connection (Figure 4), the tubing touches
the bottom of the column endfitting, with no void between them.
Figure 4: Proper Tubing/Column Connection
The presence of a void in the flow stream reduces column
performance. This can occur if a Parker ferrule is connected to a
Waters style endfitting (Figure 5).
Note: A void appears if tubing with a Parker ferrule is connected to a Waters
style column.
0.090 inches
Figure 5: Parker Ferrule in a Waters Style Endfitting
There is only one way to fix this problem: Cut the end of the tubing
with the ferrule, place a new ferrule on the tubing and make a new
connection. Before tightening the screw, make sure that the tubing
bottoms out in the endfitting of the column.
Conversely, if tubing with a Waters ferrule is connected to a
column with Parker style endfitting, the end of the tubing will
bottom out before the ferrule reaches its proper sealing position.
This will leave a gap and create a leak (Figure 6).
Note: The connection leaks if a Waters ferrule is connected to a column with a
Parker style endfitting.
0.130 inches
Figure 6: Waters Ferrule in a Parker Style Endfitting
There are two ways to fix the problem:
1. Tighten the screw a bit more. The ferrule moves forward, and
reaches the sealing surface. Do not overtighten since this may
end in breaking the screw.
2. Cut the tubing, replace the ferrule and make a new connection.
Alternatively, replace the conventional compression screw fitting
with an all-in-one PEEK™ fitting (Waters Part Number PSL613315)
that allows resetting of the ferrule depth. Another approach is to