This means that our software program cannot locate the hardware protection key required for its proper functioning. It must be connected to the computer on which the program is running.

1. The key can be one of 4 models, 3 USB models are illustrated below. If you own a beige paralllel port key, it cannot be run under Windows 10 and 11 (your only option is to update your software).

   Cyan protection key No driver installtion required	Black protection key You must install driver 7.6.8	Purple protection key You must install driver 7.6.8

   You can download the 7.6.8 driver file by clicking here.
2. There are a few reasons why the program might not be able to find the key. Here are a few things to verify or do:
   

   1) Drivers must be installed before connecting the key. If you did not or are unsure, try uninstalling it (using the same software as for installation), restarting the computer, reinstalling the drivers (try with the default settings), restarting the computer then connecting the key.

   2) Make sure the key is connected directly to one of the computer's USB port (not via a USB hub, monitor or else).

   3) Make sure you are using the right key for the software. Check that the software model it unlocks written on the key is the same as the software model you are trying to use (written in its Welcome screen at startup).

   If these suggestions does not work, contact technical support.
   

1. They can load and analyse much larger images.    
2. They are approximatively 15% faster.
3. Unfortunately they can only acquire images via TWAIN64 drivers and these are rare (as of Nov 2021 our STD4800 deos not have such drivers but our thrid generation LA2400 has). When there is no such drivers, the only option is to acquire images with the scanner manufacturer software, save the images and load them in our programs for analysis.

1. Why are there empty classes (bins) in the Root Distribution Histogram: This is related to the image pixel size and the fact that root diameters are measured as a multiple of it. All along the root, its diameter is measured and can be 1, 2, 3.... or N pixels wide. If pixel size is 0.15mm, then the root can be 1x0.15 (=0.15), 2*0.15 (=0.30), 3*0.15 (=0.45)....N*0.15 mm wide. So depending upon how your classes are divived it is possible that some never get any root in them (if your classes are 0.0 =to< 0.10 =to< 0.20 =to< 0.30 =to< 0.40 as example the first and third would always be zero). How to minimise this? 1) Turn on "Pixels interpolation" in Analysis/Measurements. Note that this works only with GREY LEVELS images. When it is active, WinRHIZO interpolates the root edge position to a fraction of a pixel. So diameter can be 2.24 pixels wide (insstead of 2 or 3) for example. 2) Set root diameter measurement mode to more precisely. In Analysis/Measurements, next to Precision, choose More or Maximum. 3) Modify your diameters classes.   
2. Why some sums of diameter class data are not equal to total values: WinRHIZO produces two volume (and surface area) data, one measured (more precise) and one estimated. The Estimated root volume is calculated from the estimated root average diameter calculated from the measured root projected area and length (using trigonometric formulas assuming roots are cylindrical). The Measured root volume is the sum of all volumes calculated along the root length using the measured diameter at these points. For a comparison of both volume measurement methods, refer to the publication below. Essentially it shows that estimated volume precision decreases when root sample diameter variations increase.

     

   Rose L. (2017) Pitfalls in Root Trait Calculations: How Ignoring Diameter Heterogeneity Can Lead to Overestimation of Functional Traits. Frontiers in Plant Science, Volume 8, Article 898, May 2017.

   The surface area and volume per diameter class of the root distribution graphic are computed precisely with the measured root diameter. For each pixel of the skeleton, the root volume and surface area are calculated with the punctual root diameter at the pixel position and added to the proper diameter class. This method is more precise than estimating the class volume and area from the class average diameter.

   You can get more information about how measurements are made in WinRHIZO Instruction Manual in Appendix C. 

3. Slow analysis: The settings/functions below have a high impact on WinRHIZO’s analysis speed.
   1) Filters in Analysis/Debris & Rough Edges Filters.
   2) The Very Pale Root option in Root & Background Detection.
   3) Using lasso or circular analysed regions instead of rectangular.
   4) Analysing a Color (instead of grey levels) image or doing a Color Analysis (in Root & Background Detection).
   5) Using Exclusion Regions.
   The computer processor and RAM memory speed have a strong impact on analysis time. The WinRHIZO version also has an impact on speed. WinRHIZO 2021 is faster than 2016 which is much faster than 2012. WinRHIZO 2021 uses mutli-threading, older versions do not. Turning on "Multi-Thread" and turning off "Data 100% identical to previous versions" increase the analysis speed significantly.
4. How do we verify that the data the software collects off of the root sample is accurate? Is there a way to verify or calibrate the system? If you bought a complete system from us you do not need to calibrate, only to use it correctly.
   1) Make sure that Scanner.cal is installed (WinRHIZO Manual Chapter 1).
   2) Scan according to our instructions (see The STD4800 or LA2400 Scanner for WinRHIZO Manual and the Video Tutorial that is in the WinrHIZO folder on your computer).
   3) Verify that roots are well detected (see the Overview & Introduction Chapters in WinRHIZO Manual); a) By looking at skeleton lines (there should be a line over each all roots) and b) the Detected Roots image (roots should be black and background white). Learning WinRHIZO using our demo images and the Introduction chapter is very important as all required notions are introduced there.
5. Can we scan multiple root systems per image? Yes. If you click the image, all of them will be analysed together as a single root system (one line of data for all of them). If you want data separately for each one, space them so that they do not touch each other before scanning and at analysis time make an image selection around each one. You will get one data line per root system.    
6. Can we cut and scan a large root system in multiple images? Yes for most analyses (root morphology and link analysis), not for some (like topology and developmental analyses which require a complete root system). If you give all subsamples the same name during analysis, their data can be merged after in XLRhizo.
7. Can WinRHIZO automatically measure roots in soil? In rare situations yes. Note that we do have two families of product for root measurement.
   1) WinRHIZO Tron which is a manual root tracing program. It is made for roots in soil or roots which do not have a good contrast with their background. It will work on any image in which you can see the roots (and make a calibration).
   2) WinRHIZO Basic, Reg, Pro or Arabidopsis. These can automatically measure washed roots or roots with a
   good contrast with their background. The Pro version (or Arabidopsis) is required for color analysis (when color is used to separate the root from their background). In some instances, the latter can be used to measure roots in soil, but for this to work the contrast must be good and/or the soil must be a different color than roots.
8. Can WinRHIZO automatically measure mycorrhizae?
   We have no experience in handling and preparing mycorrhizae. As such we cannot recommend a preparation method or say if a scanner can be used. We can say that images from cameras are suitable if the proper magnification is used (via lenses or microscope). There might be chances that a scanner could work for larger ectomycorrhizae.
   We have received images from users over time, some of which could be analysed and some which could not. The reason for the success/failure came mostly from the variation in appearance which results from the differing preparation methods. In order to measure mycorrhizae, they must contrast with the roots and background. In some cases when the root system is too dense we can only quantify the area of the image covered by mycorrhizae, roots and the surrounding background. When the root system is not too dense and a good color contrasts is achieved between root and mycorrhizae, we can measure the root morphological parameters (root length, area, volume...) for mycorrhized and non-mycorrhized roots. In the latter case, the Pro or Arabidopsis version is required because color analysis is used.
   So before recommending WinRHIZO for a mycorrhizae application we ask for a sample image taken in the real conditions of your project (an image that you plan to use for analysis).
9. Root hairs. There is no special treatment for root hairs by default (with our automatic settings). They might be included in root or background data. It depends on how their grey levels are close to root vs background and this is not uniform among species, users, image acquisition settings etc. When dealing with root hairs, a manual Root & Background Detection classification might be required so that hairs data fall into the group you want. The way to determine in which group hairs have been classified is to look at the "Detected roots" image after an analysis. You can also visualize this classification while setting up the Root & Background Detection parameters (so you can choose the right ones).
   Note that if hairs have a different color than roots, then Color Analysis might produce better results.
   The microscope is very likely required to identify and measure hairs individually. With a scanner, it is rather an envelope that can be seen and measured (maybe some individual hairs at very high dpi).
10. Nodules Nematodes. You can count nodules manually with all versions of WinRHIZO (by clicking each of them).
    If nodules are not touching each other and if they have color contrasts with the roots (naturally of via staining), then some of our programs can count them (including WinRHIZO Arabidopsis version). Note that we've rarely seen situations were it worked well automatically. There is an example of both Nodules counting methods in WinRHIZO's Instruction Manual in the "Application Examples" chapter.
11. Load Configuration not working (WinRHIZO 2021a and b). This bug has been fixed. Contact us to get the fixed version 2021c.
12. TPU light is not used. Make sure the TPU is connected to the scanner at the back of it and that you have selected "Grey levels" to the right of "Image Type" (don't choose "Color"). Important: At startup do NOT select a TWAIN Source which name begins by "WIA...").
13. Tips for a better detection of thin very pale roots. In Image/Acquisition Parameters turn "Speed priority" to off. Increase dpi (to 800 or more). If possible place roots directly on the scanner glass, aovid using water. Try the "Very pale root" option in Image/Acquisition Parameters. See the other tips in WinRHIZO Instruction manual, there is a section dedicated to this.
14. Can the WinRHIZO software analyse any image from any scanner or camera? The short answer is no. WinRHIZO is a system, made of software and hardware that were designed to fit each other. Since our scanners produce highly contrasted images, free of shadows, the analysis is very easy and fast (using WinRHIZO's default settings). If your images do not meet those criteria, then default settings cannot be used and the operatr has to experiment with different settings. The analyiss is thus more complex, difficult, time consuming and most of the time less accurate than those made on our scanners' images. The most frequent problems that we see is that not all roots are detected or that artefacts (shadows, debris...) are analysed as roots. Although there are filters to remove some of such artefacts, it is not guaranteed that they will work. Over the years we added features to allow to analyse roots from different media or imaging device. These must be tested before purchase to determine if WinRHIZO can analyse them or not. Please note also that when no hardware is purchased from us, our people at technical support won't be able to help in case of problems (in other words, we only support the hardware we sell).