Filling the Calibrant Bottle in 6200/6400/6500 Series LC/MS Systems

Answer

This Information Applies To: Agilent 6200/6400/6500 Series LC/MS Systems

Issue

Instructions on how to fill the calibrant bottle on 6200/6400/6500 Series LC/MS Instruments when calibrant levels are running low.

Background

The ability to autotune your Agilent LC/MS system relies on providing the instrument with the correct composition and concentration of calibration solution.

Items Required

The type of Calibrant used for autotune depends on the type of Ion Source installed on your Instrument. 

• Clean Glass CDS bottle (p/n 9300-2576), one is already in the instrument
• And one of the following:
  • LC/MS Calibration standard, for ESI-TOF, 100 mL (p/n G1969-85000)
  • LC/MS tuning mix, for ESI, 100 mL (p/n G2421-60001)
  • LC/MS tuning mix, for APCI/APPI, 100 mL (p/n G2432A)
  • LC/MS tuning mix for APCI, low concentration, 100 mL (p/n G1969-85010)
  • LC/MS tuning mix, for MMI, low concentration, 13 compounds, in ACN, 100 mL (p/n G1969-85020)

Steps to Follow

Caution: There is a risk of contamination when performing this procedure. Take care to avoid contamination of the tuning solution or the calibrant delivery system during the following steps. Wear clean powder-free nitrile gloves and make sure all glassware and equipment used is free from contamination.

1. From the MassHunter Acquisition Tune View, go to the Manual Tune tab, and make sure that the Calibrant on check box is cleared (Figure 1). This way, the calibrant bottle is not pressurized when you are unscrewing the bottle.
   

   
   
   Figure 1. Clearing the Calibrant on check box
   
   

2. Open the door or remove the cover on the front of the LC/MS to access the calibrant bottle. See Figure 2
   

   Caution: Some force may be required to start the unscrewing of the calibrant bottle. As the bottle is glass, avoid using excess force during removal that may damage the bottle or the bottle cap fixed to the instrument. If the bottle cannot be removed easily, then confirm that the calibrant is turned off in MassHunter, and then contact your local Agilent service representative for further advice.

3. Gently unscrew the calibrant bottle in a counterclockwise direction to remove the bottle from the LC/MS. See Figure 2.
   

   Warning: Wear appropriate Personal Protective Equipment, including gloves and safety googles when handling tuning solutions. See the specific MSDS supplied with the product for further details

   
   
   Figure 2. Removing the calibrant bottle from the LC/MS 
   
   

4. If the old calibrant remaining in the calibrant bottle is out of date or your tuning results suggest it is contaminated, discard the old tuning mix remaining in the bottle. Then rinse the calibrant bottle a minimum of two times with clean LC/MS grade Acetonitrile. If you need to flush you Calibrant Delivery System (CDS), see instructions in the following article: Flushing the calibrant system in your LC/MS Instrument.
5. Confirm that the tuning mix you are using to refill the calibrant bottle is correct for your Ion Source type, application, and mass range. Using the wrong type of tuning mix may cause contamination or will cause autotune failures.
6. Refill calibrant bottle to about 1/3 full with the appropriate tuning mix. For 6400 Series LC-QQQ systems, the tuning mix is used neat without any dilution steps. 

   Tips: 6200 Series and 6500 Series LC/MS with an Agilent Jet Stream source installed may need the tuning mix diluted if you get error messages to instruct you to dilute the calibrant, or if the tune report shows mass abundances greater than 750,000 counts. See the article Diluting the Tune Mix on 6200 TOF and 6500 Series Q-TOF Systems.

    

7. Place the intake tube into the refilled calibrant bottle as you lift the calibrant bottle into position. See Figure 3.
8. Attach the calibrant bottle onto the fixed bottle cap inside the LC/MS, then turn clockwise to tighten. See Figure 3.
9. Check that the inlet tube is submerged in the tuning solution. 
   

   Caution: The calibrant bottle only needs to be snug. Do not overtighten the bottle. However a loose and leaky bottle can prevent the bottle from pressurizing and supplying calibrant to the nebulizer, and can lead to evaporation of the contents.

   
   

   
   
   Figure 3. Reinstalling the calibrant bottle into the LC/MS 
   
   

Learn how to effectively maintain your Agilent 6200/6400/6500 Series LC/MS Instruments: LCMS-6400-1120sV2 - Tune and Calibrate Agilent 6400 Series Triple Quadrupole LC/MS with MassHunter Acquisition (Ver. 10.0) LCMS-ULTV-1120s - Tune and Calibrate Agilent Ultivo LC/TQ with MassHunter Acquisition (Ver. 1.1) e-learning courses available from Agilent education