MassChrom® 25-OH-Vitamin D3/D2 in Serum/Plasma - LC-MS/MS

MassChrom® 25-OH-Vitamin D3/D2 in Serum/Plasma - LC-MS/MS

  • 25-OH-vitamin D2
  • 25-OH-vitamin D3

Order No.: 62000/RUO, 62000/1000/RUO
200 / 1000 Tests

Suitable for every tandem-MS with sufficient sensitivity
Isotopically labelled internal standard
6PLUS1® and 3PLUS1® Multilevel Calibrator Sets
Online sample preparation
Minimal ion suppression effects

* Required Fields


This reagent kit allows for the specific analysis of 25-OH-vitamin D3 and 25-OH-vitamin D2 in serum and plasma by tandem mass spectrometry. Manual sample preparation is minimised and limited to simple and effective protein precipitation. A subsequent online trap column concentrates the analytes and separates interfering substances. A six-port-valve connects the trap column to an HPLC column which provides further purification. The use of atmospheric pressure chemical ionisation (APCI) and an isotopically labelled internal standard, plus the multilevel calibrators (6PLUS1®, 3PLUS1®) assures high performance and minimises ion suppression effects.

An upgrade for the determination of the epimeric forms of 25-OH-Vitamin D3 and 25-OH-Vitamin D2 is also available for this kit (see accessories).


Kit content 62000/RUO                                

Mobile Phase A, 1000 ml                     
Mobile Phase B, 1000 ml                      
Precipitation Reagent, 5 ml                 
Internal Standard, 40 ml                        
Rinsing Solution, 1000 ml                    
Reaction vials, 2 x 100 pcs.        


Kit content 62000/1000/RUO   

Mobile Phase A, 5 x 1000 ml                                                                   
Mobile Phase B, 5 x 1000 ml                                                                  
Precipitation Reagent, 5 x 5 ml                                                                
Internal Standard, 5 x 40 ml                                                                    
Rinsing Solution, 2 x 1000 ml                                                        
Reaction vials, 10 x 100 pcs.        
Analytical column, 1 pc.           
Trap column, 2 pcs.                   
3PLUS1® 25-OH-Vitamin D3/D2 Multilevel Serum  Calibrator Set (lyoph.), 4 x 1 ml            
MassCheck® 25-OH-Vitamin D3/D2 Serum Control, Level I (lyoph.), 5 x 1 ml
MassCheck® 25-OH-Vitamin D3/D2 Serum Control, Level II (lyoph.), 5 x 1 ml


Technical Data

Method of Analysis



25-OH-vitamin D2, 25-OH-vitamin D3

Analysis Time

5 min


The trap column and the analytical column are linked to the mass spectrometer through a 2-position 6-port valve.



Sample Preparation

  • Pipette 100 μl research sample/calibrator/MassCheck® control into a 1.5 ml reaction vial.
  • Add 25 μl Precipitation Reagent.
  • Add 200 μl Internal Standard and mix 20 s (vortex)
  • Incubate 10 min at +4 °C.
  • Centrifuge 5 min at 15000 x g.
  • Transfer 200 μl supernatant into an autosampler vial.
  • Inject 10–50 μl of the supernatant into the HPLC-MS/MS system.

Sample Stability

The prepared research samples are stable up to 4 days at room temperature and up to 1 week at +2 to +8 °C. For longer storage (maximum 2 weeks) deep-freeze the research samples below -18 °C.

Additional Info

HPLC parameters

Pump A delivers 100 % Mobile Phase A for loading the trap column.
Pump B delivers 100 % Mobile Phase A for separating analytes on the analytical column.

Injection Volume

10 to 50 μl (mass spectrometer dependent)


Pump A: 0.02–1.00 ml/min
Pump B: 0.60–1.00 ml/min


APCI positive



Additional Info

LC-MS/MS parameters

Kit Components

Components available separately

Mobile Phase A

Order no.: 62001/RUO

Mobile Phase B

Order no.: 62002/RUO

Precipitation Reagent

Order no.: 62003/RUO

Internal Standard

Order no.: 62004/RUO

Rinsing Solution

Order no.: 62009/RUO



Controls available separately