The

Sample Spotter
Model SES 3202 / IS 03
With
Platform for Chromarods

USER GUIDE

-1-
 Mounting the Chromarodplatform to the Spotter

The Sample Spotter Model SES 3202/IS 02 is a carefully prepared package for spotting
TLC-Samples on both:

- TLC Plates and

- Chromarods used with latroscan

For reasons of save transportation the system is sometimes delivered with the platform
for TLC-Plates.

For use with Chromarods in such a case you have to unscrew the platform for TLC-
Plates and replace by the platform for Chromarods. The screws are underneath and can
be unscrewed with an Allen key or appropriate screwdriver.
Please be careful, as the Instrument is a high precision mechanical device.
After mounting the Chromarod-platform, continue the procedure for mounting the syringe
and the adjustment of the chromarod-platform as described in the manual part called:

"Spotting on Chromarods with Spotter Model 3202/IS 02"

In some versions we are delivering a block of aluminium with holes as a supplement.

This piece mounted, will increase the potential stroke of the piston of the syringe.
Normally one can do without, as the concentration of the sample can be adjusted by
the correct amount of solvent. For those who want a complete stroke of 2 microliter on
the delivered syringe, have to include that block into the construction if not yet done.

ATTENTION

If not otherwise defined and ordered, we deliver the template for Chromarods to the
users of Iatroscan TLC-FID-Analysers.
If the template for spotting on plates would be wanted, we can deliver this part to
replace on customers order and invoicing the costs.

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1. The Principle

The sample is drawn up in the needle of a microliter syringe. The syringe piston moves
within the needle and comes to the tip at its lowest setting. Thus all the sample in the
needle is always ejected and residue levels are minimal since the only void consists
of the clearance between the piston and the inner wall of the needle. The dead volume
is therefore very small.
The piston is motor-driven between two mechanical stops. The electronic control unit
always detects when the piston has reached a stop and then halts the motor or reverses
the direction of motion for the rinsing process. This limits the maximum force which can
be developed and precludes any damage. Even a drive system blockage due to
incorrectly mounted syringes or other such improper procedure, cannot lead to serious
damage.
The bottom stop is defined by the syringe itself. Irrespective of the type of syringe and
the tolerances of the mounting and the drive, the piston is always moved fully in the
syringe.
The upper-stop is continuously adjustable. Any desired volume between 2 % and 100%
of the syringe volume can be set. The drive motor moves the piston via spring elements
and spindle to both stops, so that the reproduciblity of the piston stroke depends only on
the mechanical constancy of the stops, i.e. on the syringe mounting and on the mounting
of the adjustable upper stop. Both these mountings are established without intermediate
elements, by direct clamping on two precision polished steel rods, so that the
reproduciblity is better than +/- 0.05 mm, i.e. better than 1% of the syringe volume for the
usual scale length of 55 mm.
However the absolute accuracy is determined by the syringe itself and by its scale,
according to which the movable stop is set.
The electronic control unit contains the electronic power circuit for the motor, the control
logic and the function keys. Power is provided by a separate adapter which conforms to
strict electrical regulations. The adapter delivers only safe low output voltages. Syringe
directions, up and down are started by pressing the marked key " " or " ".
Movement stops when the piston reaches the stop or when the "Stop" key is pressed.
For rinsing, 2 complete piston strokes are started by pressing the key " ". These
strokes rinse with solvent or, preferably, with the next sample. Carry-over is then less
than 1%.

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A very much slower injection speed can be switched on or off with the key "L". The time
for ejection of the complete syringe volume is extended from 7 seconds to 7 minutes by
very short switch-on times during which about 1% of the syringe volume is ejected each
time, with 3 second delays between successive ejection pulses. In periods between the
ejections, the solvent can evaporate. These pulses are not consistent and therefore
cannot be used as Individual applications but only for controlling of a slower application
of a total volume set by the syringe stop. This makes it possible to produce very small
spots down to the needle diameter of 0.5 mm even with large sample volumes.
The dosing system consists of the drive spindle, the syringe mount with syringe and the
adjustable stop. It moves vertically in the spotter head. Thus the needle dependably
touches the surface of the layer, irrespective of the thickness of the TLC plate. The
spring compensates a part of the weight of the dosing system so that the contact force is
minimal. This will not cause any damage to the customarily used TLC layers.
The spotter head can be swung to the front or to the rear by the swivel levers attached
on both sides. In the front position, the sample vessel is brought to the needle and the
aspiration process is started by pressing the key " ". The sample vessel is thereby
about 50 mm in front of the plate, to rule out contamination of the TLC layer by falling
particles.
On swinging-back the dosing system is lowered until the needle tip rests on the surface
of the layer. Ejection of the sample is then started automatically by microswitch.
During each swing-forward and swing-back the spotter head is moved a fixed distance to
the right to the next spot position. The application pattern is a 8 mm pattern adapted to
distance of the Chromarods in the Chromarod-holder SD-5.

The Chromarod-holder SD-5 is placed into the adjusting frame on the whole assembly.

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2. Operating in general

2.1 Set up

Stand the unit on a flat and level surface. Remove the transportation securing
rubber ring on the spotter head. Insert the multipin plug connector of the control
unit into the socket on the rear of the main frame, and secure the connection by
turning the sleeve ring. Plug the adapter into an AC power outlet. The lamp in the
key " " lights up.

2.2 Inserting the Syringe

Swing the spotter head in oblique position with the swivel lever. Release the
milled screw in the adjustable stop and fix this at the upper stop. Run the driver
about 10 mm below the adjustable stop pressing the keys " " and "Stop". Push
the piston completely into the syringe and hook the piston plate into the driver.
Push down the syringe body such that the needle tip touches the upper surface of
the front part of TLC surface or Chromarod layer. Then tighten the milled screw
on the syringe mount.

2.3 Setting the Volume

Swing the spotter head to the front and then press the key " ". When the piston
has come to rest, read the volume against the syringe scale.
If a change is necessary, release the milled screw on the adjustable stop. By
pressing the keys " ", " ", and " Stop ", run the piston to the desired volume.
Then fix the stop position with the milled screw. Check the setting by pressing the
key " " and taking the volume reading when the piston has come to rest again.

2.4 Spotting of Samples

Swing the spotter head to the front, compress the two disengagement levers and
push the spotter head to the left stop position. Release the disengagement
levers. Load the TLC-Plate or Chromarods and position it to the front and left
against the stop. Adjust the frame so that the middle of the first Chromarod is
adjusted to the middle of the syringe-tip.
Bring the first sample vessel to the needle, such that the needle dips well into the
sample. Press the key " " to start a rinsing process. When the piston has
come to rest, ( i.e. when the lamps in the keys " " and " " have stopped
flashing) press the key " " to draw up the sample. Remove the sample vessel
into another rack to separate the already spotted samples.

-5-
 If carry-over can be tolerated or the concentrations of the samples are
similar the rinsing can be omitted. Swing the spotter head slowly to the rear.
The tip of the needle lowers on to the TLC-Plates or Chromarods and spotting
starts automatically. When it has finished, swing the spotter head to the front and
repeat the entire process with the next sample vessel. After placing the last spot,
take off the frame with the Chromarods and carry out a rinsing cycle of the
Syringe with pure solvent. After finishing work, move the driver between the stops
by pressing the keys " " and "Stop" and remove the syringe if required.
If the instrument will be used in same conditions later, the syringe can be left in its
position. Be careful so that the tip of the syringe will not be bent. Uncontrolled
touching may damage the syringe such that a replacement with new might be
necessary.
Take the electronic drive with adapter from the power outlet.
By pressing key "L" spotting may be slowed down. The lamp in this key lights.
Slow spotting mode persists until the key "L" is pressed again. Evaporation can
be intensified with a small airpump ( membrane pump ) or nitrogen stream.

3. Hints for use

The instruments was carefully adjusted and checked before delivery. Rough
treatment and dirt may disturb the operation. Please check the following functions
from time to time:

The syringe should not stick while slowly lowered onto the absorbing layer. It should
be easily movable in vertical direction.

The motor should run smoothly between stops, even with maximal volume setting.

Keep the sliding surface - two horizontal and two vertical round stainless steel rods -
Clean, lubricate them at times with a very thin film of acid-free and carbon free
lubricate.

Syringe pistons should be easy to move an syringes free of leakage.

Syringe needles are often not straight. Therefore the spotting position depends from
the orientation of the syringe in its mount. Always keep the same orientation - scale to
front - when mounting a syringe. Then the spotting array is constant from Chromarod
to Chromarod.

A non-defined amount of sample sticks to the needle tip after filling the syringe. It may
be 20 nl with a 1 l syringe. You get the optimum reproducibility by wiping the needle
from the syringe to the tip with soft paper.

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4. Technical Data

Contact Force 1 N max.

Reproducibility of Delivered Volume 0.5 %
Reproducibility of Positioning 0,2 mm
Rate of Delivery 100 nl/s
(1 l Syringe)
In "L"-mode 15 nl/s
Power 220 V, 50-60 Hz, 5 VA
Or 110 V, 50-60 Hz, 5 VA

Dimensions (w * h * d) 330 * 340 * 260 mm

Packed 430 * 420 * 370 mm
Weight 5,5 kg
Packed 7,5 kg

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 SPOTTING ON CHROMARODS WITH SPOTTER MODEL 3202 / IS-03

ATTENTION PLEASE !!

The application of the samples onto the Chromarods is one of the most important
elements for quantitative analysis with the Iatroscan.

The quality of the quantitative result is directly related to the quality of the sample
spotting process.

There are a number of points to pay attention to, as far as the fixing of the syringe and
the alignment of the syringe to the Chromarods are concerned.

We recommend following procedure:

1. First, move the syringe-mount by means of the swivel-lever to its 45 degree

inclined position in order to fix the syringe. Fix the syringe provisionally with the
fixing screw on the syringe mounting part of the spotter. Take measurement with
the tip of the needle to the level of the rods, first without touching.
The fixing of the syringe has to be done very carefully and with precision. The
position of the syringe is important to avoid pressure of the needle on the
Chromarods. To high pressure of the needle on the Chromarods would damage
the layer or even break the rods. Failure to make contact between needle and
Chromarods, however, would prevent sample spotting taking place or result in
incomplete spotting of sample. This would effect the accuracy of the analysis.

2. Place the Chromarod-holder SD-5 with Chromarods on the platform, such that
Chromarods fall into the grooves of the mechanical fixing device. Now move the
movable part of the platform in vertical direction to define the distance of spotting
on the rods and fix this direction by means of the related screws.

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 3. Horizontal adjustment.

Now move the horizontal metal parts with the grooves such that the Chromarods
are exactly under the needle of the syringe. Please be careful to move both metal
parts at upper and lower end of the Chromarods by little bits and simultaneously.
Be careful not to break the Chromarods. The correct position of the Chromarods in
horizontal direction is reached by moving the Chromarod-holder with the rods
laying in the metals with grooves.

4. Place, move and fix the syringe. The mounting of the syringe. Is a carefully and
smoothly executed trying until the needle has its exact position. The needle should
be exactly central to the Chromarods the pressure on the rods must be sufficient
light. If the syringe is mounted exactly, the light spring on the moving part of the
spotting head prevents the excessive pressure on the rods. Before spotting the
samples, it is necessary to check if all the spotting-positions on the individual
Chromarods are in contact with the spotting needle as described above.

Thus the correct position of the syringe is achieved when the needle lightly
contacts the central point on all the rods at the desired sample spotting location.

If the right position is achieved, fix all screws of the mechanical adjustment. If
correctly done, there will be no need of adjustment for a long time any more. The
Chromarods will remain in stable position for spotting in future.

So, with the syringe mount fixing-screw and the adjustable upper stop, the
pressure of the needle on the Chromarod and the spotting volume of sample can
be adjusted respectively.

This procedure should be performed and verified before commencing to spot the
actual samples.

5. Sample Spotting

Taking the sample into the syringe and spotting it is a function as described in the
operating instructions, similar to spotting on TLC-Plates as described under
"Working Principle."

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6. The speed of the sample spotting can be regulated by turning the Potentiometer
with "L"-Position switched on the electronic motor-control. The type of solvent for
the sample and the spotting speed should be adjusted so that a balance is
established between the sample absorption rate on the TLC coated rods and the
evaporation of the solvent. In this manner, we reach the best conditions for a
complete absorption of the sample volume and the smallest dimension of the
spot-dispersion on the Chromarods. This is important, the smaller the spot, the
better will be the resolution of the peaks in the chromatogram after development in
a suitable eluent mixture.

7. After the spotting of the last sample the syringe is rinsed out immediately a couple
of times in order to prevent carry over of sample. All methods and procedures
applicable to the working and cleanliness in avoiding sample carry over are similar
to those observed in classical plate TLC.

8. After the samples have been spotted carefully and dried in a drying-oven, the
Chromarods are placed, together with the Chromarod-holder, into the
development chamber.

It is Important to provide a pre-saturated atmosphere of the eluent mixture by

means of an inert filter paper of appropriate size. This avoids development
speed variations from rod to rod, occurring as caused by differences in vapour
pressure within the chamber. The reproducibility of the chromatograms will
depend upon this control.

9. After the desired development time, the rods are taken out of the chamber.
Please pay attention not to contaminate the developed rods with air dust or
other particles in the air while transporting!

After eventual drying in an oven, the rods in the rod-holder can be placed
into the latroscan for scanning.

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 Analysesysteme
Friedhoftstrasse 7-9 - 55234 Bechenheim / Germany
Tel: +49 (0) 6736 1301 - Fax: +49 (0) 6736 1305 - Email: ses_analysesysteme@t-online.de

2.0

2.1
2.2

2.3 1.2
1.1
1.0
2.5
2.4
1.3
2.6

2.7

3.1
3.2
3.0 3.3

1.4
3.5 3.6
1.5 3.4

1.0 Main Frame 2.0 Dispensing System 3.0 Electronic Control Unit

1.1 Template 2.1 Drive Spindle 3.1 "" Rinse Cycle

1.2 Thumb Screw 2.2 Spotting Head 3.2 " L " Pulse Spotting Mode
1.3 Swivel Lever 2.3 Disengagement Lever 3.3 " Stop " Terminate Motion
1.4 Plate Stop 2.4 Adjustable Stop 3.4 "" Syringe Empty
1.5 Front Strip 2.5 Driver 3.5 "" Syringe Fill
2.6 Syringe 3.6 Connection Cable
2.7 Syringe Mount

high high

low low

P2
waiting Time
P1
Spotting Time
(between Spotting)