How to make a calibration curve and calculate sample concentrations using Excel – Video Tutorial

How to make a calibration curve and calculate sample concentrations using Excel - Video Tutorial

Excel Calibration Curve Video TutorialWorking in the laboratory, there are a number of different ways that we can calculate the amount of an analyte present in a sample by comparing them to standards. You could use a single external standard, a calibration curve,  internal standard or use standard addition. Learning to use these different methods of calculation is essential to working in the laboratory and is a essential part of any laboratory training program.

This is a video tutorial for making an Excel sheet to create a calibration curve using six standards and using it to automatically back calculating unknown sample concentrations.

The sheet also includes a dilutions factor calculator using which the concentration of analyte in the undiluted samples can also be automatically calculated.

We also show you how to protect the Excel sheet so that the formulas cannot be altered by mistake and also you can further validate the Excel sheet for use in a regulated environment.

For best results see the video in HD, in full screen mode and use headphones for better sound clarity.

From the video tutorial you will learn how to –

  1. Enter data for a calibration curve in excel
  2. Make a calibration curve with linear regression trend line
  3. Display the linear regression equation of the line on the graph
  4. Display rvalue of the regression line on the graph
  5. Calculate the slope and intercept for the regression line using formulas
  6. Calculate the concentration of unknown samples using the equation y = mx + c
  7. Calculate dilution factor for samples which are diluted prior to analysis
  8. Calculate undiluted sample concentrations
  9. Protecting the excel sheet so you can validate it in the future
  10. Formatting the excel sheet so it can be printed properly on one sheet of paper

What can you use this sheet for?

You can use this sheet for calculating sample concentration from a standard calibration curve for any technique like HPLC, GC, UV, AAS or any other technique where linear regression is used. You can also use it in method validation to evaluate linearity of the response and establish the range of the method.

Please leave your comments below!

Please share your feedback on this video and let us know what other topics you would like us to cover in the future by leaving your comments below and don’t forget to share it with your friends!

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  1. Thank you very much for this nice video.
    Actually I am interested in knowing how can I calculate and represent in the chart the error of the result. When I calculate for instance a concentration by means of a calibration curve, I got a value. But I need to know how good is this value and a +/- around this value.
    Thank you,

    1. I would like to thank you for this excellent video. I WOUNDER HOW I CAN COPY THE VIDEO SO I WOULD BE ABLE TO WATCH IT AGAIN IN CASE I LOST CONNECTION.

      1. Hi, I am glad you liked the video, we do not have an option for downloading the video currently. we will check and see if it can be done.

        1. Hello Mr. Arora
          thanks a lot for uploading such a useful video….I also want to upload this video as it is very useful to the students who face the problem to prepare calibration curve in HPLC system software.

          Thanks and God bless you.


          1. Hi Dr. Sahrma, let me know which site you want to upload it on and we will send you the link and permission. Glad you liked it!

    2. Thank you for nice video. I have small question. Do I need to prepare calibration curve each time, conduct analysis. Or I can prepare once and use it for a couple of times. For example I run analysis of nitrate and get r2 0.998 to day, am I required to prepare calibration curve tomorrow for the same parameter

      1. My advise is to prepare a calibration curve every time you conduct the analysis as the operational parameters and instrument performance can vary day to day. If this is not possible every time then at least run standard injections in between sample injections.

  2. Just wanted to express my gratitude at you uploading this clear and helpful video that has aided me in determining Sodium Nitrite concentrations, couldn’t have done it with out you.

    Best regards

  3. Any clue to calculate and represent the error of a calibration curve?

    1. Hi Fidor, the regression line that we make using the calibration data accommodates for the errors. We usually look at the r square value and test for non zero slope to evaluate the suitability of the calibration curve. The errors for the individual points can also be shown if we have replicates (minimum 3) for each of the calibration points, then we could add error bars to the values. Also there is a method to produce a 90 or 95% confidence regression line for the curve. These are all statistical methods, how ever in analytical applications the calibration range is thoroughly evaluated for accuracy and precision during method validation. I hope my longish answer makes some sense!

  4. Hi, Thank you for this useful video!I have question: how do you calculate the concentration of your samples when the calibrator concentrations fit a sigmoidal curve?Is the process similar to what you showed in this video?

  5. Hi, Thank you very much, I am glad to see the video, so much helpful for me , Could I downnloading the video currently ? , Thanks

    1. Hi,
      I understand you have difficulty downloading the video.Please let me know which video you are referring to so that we may offer help

  6. very informative video. i would be grateful if you demonstrate how to calculate drug content in tablet using calibration curve .thank you

  7. This video has been very useful to me, thanks very much for your work.

  8. Hi. Thank you for sharing your knowledge with us, it is very informative. Could you do me a favour by sending this video (How to make a calibration curve and calculate sample concentrations using Excel – Video Tutorial) to my email? I want to download it but not able to. Thank you very much in advance.

    1. Hi Syazana, It is a big video, about 800 MB will be difficult to mail it. I am glad you liked it, please feel free to refer to the site any time!

  9. Hello Dr. Saurabh Arora sir,

    thank you so much for sharing very informative video with us regarding how to prepare a calulator on excel sheet.

    abhishek aggarwal
    M.Pharma ,PGDPRA

  10. thank you very much Dr. Saurabh Arora for sharing. it is very informative and helpful to me. Can you show us how you calculate inflection point from S- shape curve using excel? and thank you again.


  11. Thank you very much Dr. Saurabh Arora for this, I am studying drug release and need to make dilutions of the aliquots I take out from dissolution at each time point. Like say for example I took 5 mL and then dilute it 1 in 100 to be able to read absorbance, how will I calculate the undiluted concentraion in that case.

    Looking forward for your reply.

    Thank you


  12. This is such a good demonstration of how to produce a calibration curve in excel. Thank you for taking the time to do this. It is really helpful to me and I am sure to many others.

  13. Thank you so much. I found this very useful.
    I do have a question though. When I referenced the cell with a link to (x-intercept value/slope value) in order to calculate other concentrations the answer was different than when I manually typed it in. The difference was slight (e.g 39.4 vs 39.2). In each case the referenced values were the same, the only difference being one had the intercept/slope values manually typed in and the other had a link to the cells which in themselves had a formual to create the intercept and slope values. Why would this be?

  14. wooooow, you have made my working so simple for me. thank you very much. West Africa (Ghana) appreciates.

  15. merci beaucoup pour la video et pour les explications ,c’est très instructif et explicite
    je voudrais si vous le permettez de compléter par toutes les utilisations de l’excell pour la realisation des validation
    thank you very much i hope that you undersand french

  16. Can you tell me why you changed the concentration value of 15 to 12 before inserting the intercept formula?

  17. Very nice and useful video. It would be nice if you could stress the laboratory analysts on the importance of checking the standards at periodical intervals such that the response from the instrument is within the permissible limits of error and integrity of the standards is ensured. You may get a good r value, but the instrument response for the standards may be low. How about advocating having check samples with known value.

    1. Thank you for your appreciation and I also share the value and stress you place on the intermediate checks of standards. If you can write a short article on this topic with your experiences we will be happy to publish it with you as the author.

  18. I want to thank you so much for this video, its so helpful. God bless you.

    1. Hi Auwalu,
      Nice to hear that. Hope now you will be able to complete your HPLC programme and earn the certificate also.

  19. Very informative.. COuld you please tell me the unit of the concentration calculated ? I mean whether it is ug/mL r mg/mL ??

  20. i want to know about when we prepare calibration curve some time regression equation show y = 0.0318x + 0.0065 and some time show 0.0984x-0.3422 so please tell me what is the difference between these equations

  21. Thank you for the video. It is clear and easy to follow. I just have one question in terms of using the dilution factor. If one has a stock solution of 6 analytes of 2500mg/L, then makes 6 standards by taking from the stock 4ml,20ml,,40ml,200ml,300ml and 400ml and making each to the mark of 1000ml; does a dilution factor play a role in final concentrations and how does one calculate for that?It doesn’t seem to make sense to me to follow the same calculation as in the template (that’s more for serial dilutions?), then again maybe I am wrong. Please explain or refer me to relevant text. Here one would be taking each of those volume from the 2500mg/L stock and making each of those volumes up to another litre.

  22. Thank you Arora sir giving me information,how to create linearity graph in excel sheet and u r excellence sir

  23. To be honest , it is very useful website and thank you for sharing your knowledge and experience.
    thanks again and we are waiting for more.

    best wishes.

  24. Hi
    Very usuful video but i have small quetion
    Please explain defference between RSD caleculation and sample concentration….?

    1. Sample concentration is the amount of analyte present in your sample. On the other hand RSD relates to the linearity of the calibration plot which you obtain a plot using 5-6 different known standard concentrations. A value of 1.00 RSD implies perfect linearity of plot and any value lower than 1.00 means slight deviation from linearity.

  25. Hi
    I appreciate you, thanks for the video. it make easy understanding
    thanks a lot

  26. helo sir, useful video for students, could you please upload the finding unknownn concentrations in dissolution studies of combination drugs.
    for combination drugs 2standard curves are prepared, so which standard curve i consider for finding unknown concentraion of mixture of drugs.

  27. The video was very insightful.
    Thank so much for sharing
    I have loads of sample that I analyzed using the spec but I have not been able to convert it fro absorbable to conc.
    This video really helped me
    But still not clear on the dilution factor calc

  28. Thank you very much for this video, indeed it clarified all my doubts. Marking it in bookmarks :)


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