Set-up and familiarization for the Wescore 5520

Vapro System Description

The Vapro osmometer is an advanced electronic adaptation of the hygrometric method of vapor pressure determination. The sensitive thermocouple and sophisticated electronics provide the means to measure the dew point temperature depression of a specimen with resolution to 0.00031 °C. Vapor pressure and freezing point are among the colligative properties of a solution. Compared with pure solvent, these properties are altered in proportion to the number of solute particles dissolved in each kilogram of solvent (water in the case of biological solutions). Thus, measuring either property is an indirect means of determining solution concentration or osmolality. The chief advantage of the vapor pressure method is that it does not require alteration of the physical state of the specimen.

Benefits include:

• 10 microliter sample size.
• Routine operation on micro samples of any biological solution, including whole blood, serum, plasma, urine, and sweat, as well as complex specimens such as tissue samples.
• None of the measurement artifacts that arise in freezing point depression measurements due to elevated viscosity, particulate matter, inhomogeneities, or other physical characteristics of the sample.Superior reliability because the measurement involves minimal mechanical complexity.

NOTE:  Vapro displays in Standard International (SI) units: mmol/kg.

How it Works

A 10 microliter specimen is aspirated into a micropipettor tip. The specimen is then inoculated into a solute-free paper disc in the sample holder, whereupon the sample holder is pushed into the instrument and the sample chamber is locked. Locking initiates the automatic measurement sequence. The sensing element is a fine-wire thermocouple hygrometer. This is suspended in a unique, all-metal mount, which when joined with the sample holder, forms a small chamber enclosing the specimen. As vapor pressure equilibrates in the chamber airspace, the thermocouple senses the ambient temperature of the air, thus establishing the reference point for the measurement. Under electronic control, the thermocouple then seeks the dew point temperature within the enclosed space, giving an output proportional to the differential temperature. The difference between the ambient temperature and the dew point temperature is the dew point temperature depression–an explicit function of solution vapor pressure. Dew point temperature depression is measured with a resolution of0.00031 °C. The microprocessor-controlled measurement cycle requires 80 seconds.

Pipette Guide
Aligns and steadies the pipette for precise application of the specimen onto the sample disc in the sample holder.

INSTRUMENT REAR PANEL

Signal Chime
Sounds a short chime at the conclusion of each measurement cycle and a long chime when certain fault conditions exist. Turn the shutter wheel to adjust volume.

Setting Up the Osmometer

Place the osmometer on a suitable work surface.

NOTE:
Avoid locations where instrument precision will be altered by thermal gradients or rapid temperature changes caused by heavy foot traffic, air vents, blowers, heaters, or windows.

Connect the power cord to an electrical outlet that matches the voltage selected on the rear panel. Avoid power circuits that are shared by centrifuges, air conditioners, or other power equipment. We recommend that you use a power line surge protector to isolate the osmometer from spikes and surges.

Verify that the sample holder is in the measurement position (sample slide is pushed completely into the instrument until it stops).

NOTE:
If you open the chamber lever before the end of the initialization cycle, the warning chime sounds.
The initialization cycle establishes the reference point for the instrument. When the initialization cycle is complete, the display screen will appear as shown on the left.

NOTE:
If you open the chamber lever before the end of the initialization cycle, the warning chime sounds.
The initialization cycle establishes the reference point for the instrument. When the initialization cycle is complete, the display screen will appear as shown on the left

NOTE:
Even though the screen at this point indicates the instrument is ready, calibration will not be stable until the instrument reaches thermal equilibration (see below).

Temperature Drift Scale
Osmolality determination involves the measurement of extremely small temperature differentials. The osmometer is thus sensitive to ambient temperature changes, which induce internal temperature changes.

While the instrument compensates for small changesthat occur over time, moving the instrument to a different area or exposing it to too much air circulation will shift the osmometer’s reading and calibration points. The Temperature Drift scale allows you to determine when internal temperature has stabilized. The Temperature Drift scale appears on the screen whenever the instrument is in the “standby” mode, and it has completed two cycles. The instrument is considered stable and ready to operate unless the indicator is against the + or - marks on the scale, indicating a changing internal temperature that can affect instrument calibration. See Note below:

NOTE:
It is normal for the osmometer to undergo a significant temperature drift during the first few minutes of equilibration. The time required to achieve temperature stability depends on the initial instrument temperature, (typically 10 to 30 minutes) but may be longer if inital temperature varies more than 5 degrees from room temperature.

NOTE:
Under normal circumstances, leave power on to keep the instrument in a ready state and to maintain stability.

Menu Selections

Two main menus (Mode and Function) are used to select modes and functions of the osmometer. Modes control how the osmometer processes samples and displays results. Use the Mode Menu to select the mode before loading a sample, or immediately after running a prior sample but before opening =the sample chamber. After you select a mode, all samples are processed in that mode until you select a different mode. Functions are specific actions the osmometer can perform. The Function Menu is not available until you have assayed a sample.

Press SELECT to move the selection arrow on the display.

Press ENTER to activate the selected menu item. If you continue to press SELECT the arrow will return to the top of the menu

MODE MENU

The Mode Menu appears upon opening the chamber while the instrument is in the “standby” mode (the instrument has cycled on an empty chamber), or when you select Mode Menu from the Function Menu, or when you press SELECT while the chamber is open.

When you activate the Mode Menu, the arrow points to the current mode. When you close the chamber, or press ENTER with the chamber open, the instrument executes the sample measurement in the selected mode. You can change the mode at any time before the measurement cycle begins, or on the last sample before the sample chamber is opened. To activate the Mode Menu after a sample has been run, either select Mode Menu from the Function Menu or open the chamber and press SELECT.

The modes are described below.

Normal Mode
For routine running of single samples. Does not display any statistical data. This mode is the default
setting upon power-up of the instrument.

Average Mode
Will run single samples while maintaining statistical data on up to 32 samples. These data include the number of samples run (1 to 32), mean, standard deviation, highest result and lowest result.

NOTE:
If a 33rd sample is run, the result will displace the first sample, a 34th sample will displace the second sample, and so on. The statistical evaluation will always be based upon the latest 32 samples, if more than 32 samples are assayed.

Average Mode is useful when you need the best possible accuracy. When the instrument is calibrated in Average Mode, calibration is adjusted to the mean value of the assayed samples. Calibration resets the operation mode to Normal Mode (see Section 3.5).

NOTE:
We recommend calibrating in Average Mode using 3 or 4 samples of each calibrating solution value.

To restart a new set of precision values, bring up the Mode Menu (the arrow should be pointing at Average Mode). The next sample assayed will be sample #1 in the data group. Or, select the Mode Menu immediately after running a sample, before opening the chamber. Select Average Mode and press ENTER. The last sample assayed will be #1 in the data group.

Auto Repeat Mode
Checks the repeatability of the osmometer on the same sample. The instrument will automatically run
10 consecutive measurements on the sample, (usually a 1000 mmol/kg Optimol sample) and display the statistical data. Since the chamber is not opened between measurements as in other modes, a short delay occurs between each measurement while water evaporates from the thermocouple. During this time the display screen shows “Sensor Drying.” Low osmolality samples (below 200 mmol/kg) may show a difference between the first and any subsequent readings if the chamber is contaminated
The Auto Repeat sequence can be interrupted at any point simply by opening the sample chamber

Process Delay Mode
Complex samples (such as leaf and other samples from which water may not readily evaporate) require long periods to reach vapor equilibrium. Process Delay Mode delays the measurement cycle after you close the chamber until you press ENTER. In research applications, this lets you delay measurement until vapor equilibrium is achieved. The measurement can be repeated without opening the chamber and to avoid vapor loss. See Appendix D for more information.

To display this menu, press SELECT after a sample has been assayed, and before opening the sample
chamber. The osmometer performs the function indicated by the arrow when you press ENTER

Function menu

To display this menu, press SELECT after a sample has been assayed, and before opening the sample chamber. The osmometer performs the function indicated by the arrow when you press
ENTER

Calibrate Function
Use this function to calibrate the instrument using the 290, 1000, and 100 mmol/kg calibration standard. Always begin with the 290 set point, then follow with the 1000 and 100 mmol/kg. See Section 3.5 for details.

Clean Test
The Clean Test consists of two consecutive sample assays on a 100 mmol/kg standard solution. The difference between the first and second assay indicates the degree of contamination in the sample chamber.

Run this test if you notice significant changes in the 100 mmol/kg calibration level.

Always use the Clean Test to check thermocouple cleanliness before assaying samples which require good linearity and accuracy in the low range. After thermocouple cleaning, use the Clean Test to verify the effectiveness of the cleaning.

We recommend that you perform this test on a routinebasis before each osmometer use session. This will allow you to monitor the condition of the thermocouple sensor and the rate at which contamination builds up in the sample chamber.

Delete Result
Deletes the last result from the data group. You can delete multiple results using this function. This function can only be used while operating in Average Mode.

Returning to Mode Menu
Press ENTER while the arrow points at MODE MENU to exit the Function Menu and return to the Mode Menu.

NOTE:
Calibration is a critical element of instrument accuracy. While it is not necessary to calibrate the osmometer while familiarizing yourself with it, you should check calibration before you assay sample material .

Sample Loading Procedure

1. Rotate the sample chamber lever upward and pull the sample slide out from the instrument until it comes to a stop, bringing the sample holder directly under the pipettor guide.
   
2. Use the forceps supplied with the instrument to place a single sample disc in the central depression of the sample holder. Make sure you have picked up only a single disc. If necessary, use the forceps and a teasing needle to separate discs. If two discs stick together, the reading will be slightly elevated. Reject imperfect discs or any that do not lie flat.
   
3. With a clean tip installed, aspirate a sample into the micropipettor by depressing the plunger to the stop, immersing the tip, and gently releasing the plunger.

NOTE:
Normally, sample droplets will not cling to the outside of the tip. If they do, they can usually be removed simply by dragging the tip against the lip of the container as you remove it. Occasionally a clinging droplet may have to be removed with a tissue, but be very careful not to wick solution out of the tip.

1. With the pipettor tip resting in the notch of the pipettor guide, position the tip about 5 millimeters above the center of the sample disc
   
2. Smoothly depress the micropipettor plunger to the stop. The specimen may drop onto the sample disc. Whether the sample droplet falls onto the disc or clings to the tip, you must complete Step 6.
   
   
   
   
   
   
   
   
   
   
3. With the plunger still held against the stop, lightly touch the micropipettor tip to the sample disc, then lift it away. The tip must briefly contact the sample disc to press it flat against the holder. The paper disc should appear fully saturated, with a slight liquid meniscus on its surface.
   
4. With the pipettor tip resting in the notch of the pipettor guide, position the tip about 5 millimeters above the center of the sample disc.
   
5. Smoothly depress the micropipettor plunger to the stop. The specimen may drop onto the sample disc. Whether the sample droplet falls onto the disc or clings to the tip, you must complete Step 6.With the plunger still held against the stop,
   
6. lightly touch the micropipettor tip to the sample disc, then lift it away  The tip must briefly contact the sample disc to press it flat against the holder. The paper disc should appear fully saturated, with a slight liquid meniscus on its surface.
   
   
7. Gently push the sample slide into the instrument until it stops. (Never close the chamber unless the sample holder is in this position.)
   
   
   
   
   
   
   
   
   
   
   
8. Grasping the sample chamber lever between thumb and forefinger, rotate it smoothly to the closed position. Closing the lever starts the measurement cycle. The display screen shows “In Process” and counts down the remaining time:
   
   When the measurement is completed, the chimesounds. The display screen shows the osmolality of
   the specimen:
   

NOTE:
Since the sample may concentrate slightly before the chamber is sealed, steps 5 through 8 should be performed with consistent timing. A warning chime sounds If the chamber is left open longer than 2 minutes. Closing the lever starts the measurement cycle. The display screen shows “In Process” and counts down the \remaining time: When the measurement is completed, the chime sounds. The display screen shows the osmolality of the specimen:

CAUTION!
Never allow the micropipettor tip, sample material, or the wet disc to touch the outer surface of the sample holder. If this occurs, abort the measurement and wipe the sample holder clean before proceeding.

CAUTION!
Break any air bubbles on the sample disc before proceeding. A bubble bursting inside the sample chamber will contaminate the thermocouple.

Cleaning the Sample Holder

To clean the sample holder and prepare for another sample:

• Smoothly rotate the chamber lever to the open position, then withdraw the sample slide.
  
• Using a lint-free tissue (not facial tissue) or acotton swab, carefully remove the wet disc andany traces of residual liquid from the sample holder.

CAUTION! Never use metal forceps to rem ove wet discs; this can damage the surface of the sample holder.

• Leave no visible residue on the holder surface. If needed, use a tissue or swab moistened with deionized water. Always clean the sample hold er with a fresh tissue or swab to avoid contamination.Avoid touching the sample holder with bare fingers.

The sample holder should appear bright, shiny, and perfectly dry before loading the next specimen.

NOTE:
Regularly perform the Clean Test using these instructions.

The Clean Test is a diagnostic feature that compares two consecutive sample assays and uses the difference to determine the contamination level of the thermocouple.

When To Run the Clean Test

We recommend running the Clean Test before each session of osmometer use, after calibration, or anytime you notice a significant (10 mmol/kg or more) shift in the 100 mmol/kg calibration. Check thermocouple cleanliness before assaying samples that require good linearity and accuracy in the low range.

After cleaning the thermocouple, use the Clean Test to verify the effectiveness of the cleaning.

Instructions

1. Run a 100 mmol/kg standard sample in Normal Mode. Observe the reading. Press CALIBRATE.
   
2. Before opening the sample chamber, press SELECT to reveal the Function Menu. Press SELECT again to point the selection arrow at CLEAN TEST.
   
3. Press ENTER. The instrument performs a second assay of the loaded sample and displays the difference between the first and second assay in approximately 2 to 3 minutes.
   
   
4. Press ENTER. The instrument performs a second assay of the loaded sample and displays the difference between the first and second assay in approximately 2 to 3 minutes.

NOTE:
If the thermocouple fails to dry within 4 minutes, the instrument will report “Check Thermocouple Head.” This indicates the presence of a contaminant (a gross contamination or fiber) on the thermocouple that is holding vapor.

If the displayed contamination level is greater than 10, you should perform the thermocouple cleaning procedures found in Section 4.

Calibrating the osmometer

Standard calibration

1. Run a 290 mmol/kg standard.
   If the osmometer reads within ± 3 mmol/kg of the standard (287 to 293), it is within acceptable calibration limits. In that case, skip to Step 4. If calibration is needed, proceed to Step 2.
   
2. With the chamber still closed, press SELECT to reveal the Function Menu. The selector arrow should be pointing at Calibrate.
   
3. Press ENTER. The instrument calibrates to the standard.
   
4. Repeat this sequence using the 1000 mmol/kg and 100 mmol/kg standards to establish baseline calibration for these standards. If the reading is not within ± 3 of the standard value, perform steps 2 and 3.

Calibration Method for Maximum Calibration Accuracy

When you need maximum accuracy, run the following calibration sequence.

1. Select Average from the Mode Menu.
   
2. Run three consecutive assays using 290 mmol/kg standard.
   
3. Select CALIBRATE and press ENTER.
   
   The instrument calibrates on the average of the three samples. This method can also be used for 100 and 1000 mmol/kg standards.

Standby or Waiting Periods

When the instrument is not in use, leave the cleaned sample holder empty and locked in the measurement position. If the chamber is left open for longer than 2 minutes, a warning chime sounds.

When in the standby mode, the Vapro osmometer is not idle. It continuously monitors its internal operating temperature and compensates for changing ambient temperatures that would otherwise result in calibration changes. It also maintains a continuous balance in its thermocouple control circuitry to ensure convergence of the thermocouple to the precise dew point temperature during the measurement cycle.

These internal functions are necessary to maintain accurate performance. That is why we recommend that the osmometer be left under power when not in use. It is also why long measurement sessions should be interrupted periodically to allow the instrument one full measurement cycle on a dry, empty chamber.

NOTE:Occasionally, after a series of runs, an osmolality reading appears on the screen after cycling on an empty chamber. This may be due to residual moisture on the sample holder. If this occurs, withdraw the sample slide and thoroughly clean the sample holder using lint-free tissue. Then return the slide to the measurement position and close the sample chamber.