Retention time standard for HPLC instrument calibration

RePLiCal comprises of unique, easy to detect peptides with selected chemical properties.

  • High number of calibrant points, covering a broad range of RTs
  • Stable RTs over long analysis times, allowing the prediction of RTs for other peptides in a timely manner
  • Includes two highly hydrophilic peptides for testing of column trapping performance
Order Number Product Package size Application Price (€)  
PQ-CS-1560 RePLiCal – complete protein, lyophilized 1 µg 250-500 injections 189.00
PQ-CS-1561 RePLiCal – peptide mix, lyophilized, digested 1 µg 250-500 injections 199.00
PQ-CS-2560 RePLiCal – complete protein, 13C,15N Lys/Arg-labelled, lyophilized 1 µg 250-500 injections 209.00
PQ-CS-2561 RePLiCal – peptide mix, 13C,15N Lys/Arg-labelled, lyophilized, digested 1 µg 250-500 injections 219.00

RePLiCals peptides have a high degree of linearity and thus can be used to predict the retention times of the peptides on a new gradient. By calculating iRTs, retention time windows can be adjusted for different gradients.

This tutorial demonstrates how RePLiCal can be used to re-calculate retention times using Skyline.

Order Information

You can place your RePLiCal order here or return the completed order form. You can also contact us directly through:


Phone: +49 (0)9405 96999 10

or Fax: +49 (0)9405 96999 28

Our prices are net prices, i.e. they do not contain value added taxes or customs fees. These taxes have to be paid by the customer.
Ordered products are usually shipped 1-2 business days after receiving the order.
The delivery costs have to be paid by the customer.

Product Information

RePLiCal is a HPLC calibration standard that is buildt from 27 lysine-terminating calibrant peptides. The peptides do not contain methionine, tryptophan, cysteine, and N-terminal glutamine residues. Also aspartic acid-proline, asparagine-proline and asparagine-glycine motifs are absent. It can be applied to calibration of LC gradients and nano uHPLC instruments using trap-elute configurations, real-time scheduling of selected reaction monitoring (SRM) experiments and recalibration of the m/z scale of mass spectrometry (MS) data post-acquisition. Find more information in the original publication.

Elution profile of RePLiCal (1 µl of a 0.5µmol/l dilution 0.5% acetonitile/0.5% TFA).  
Separation was performed on Zorbax 300SB-C18 (5×0.075mm, 3.5µm p. s.) with gradient elution at 350 nl/min from 8%-38% B (solvent A: 5% acetonitrile/0.1% formic acid in water, solvent B: 0.1% formic acid in acetonitrile. Total run time was 60 min.
Survey spectra were recorded in a range from m/z 300 to m/z 1600 using an Orbitrap XL.
RTs of RePLiCal peptides on different length LC gradients (3-40% 0.1% formic acid in acetonitrile) are compared using the 30 min gradient as a reference.
Triplicate measurements; error bars: ±2 standard deviations. High confidence in linearity is also given for very long gradients (even for 480 min).


1 µg RePLiCal or 25510 fmol (digested or undigested) equals 250 – 500 injections (50 – 100 fmol/injection).

Watch our tutorial to learn “How to use the RePLiCal RT standard to adapt for RT shifts in Skyline”.



  • RePLiCal: A QconCAT Protein for Retention Time Standardization in Proteomics Studies. Holman S. et al., J Proteome Res. 2016 Mar 4;15(3):1090-102. [PubMed]
  • Platform Independent Protein-Based Cell-Of-Origin Subtyping of Diffuse Large B-cell Lymphoma in Formalin-Fixed Paraffin-Embedded Tissue. Reinders J., et al., Sci Rep. 2020 May 12;10(1):7876. [PubMed]

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