iFlow error in opening FCS and LMD files
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Anna Dvorkin ▴ 40
@anna-dvorkin-4204
Last seen 7.6 years ago
Dear All, I have been trying to analyze files I obtained from a flow cytometer. In order to do that I am trying to use the iFlow package. After I write the following: *library(iFlow)* A GUI window opens and I choose the files I want to analyse. However, after I click on "Open" in the GUI, I receive the following message: *"Error: '.SigArgs' is shorter than '.SigLength' says it should be"* After I close that window, I can see, that the following packages were successfully loaded: *Loading required package: RGtk2 Loading required package: flowViz Loading required package: lattice KernSmooth 2.23 loaded Copyright M. P. Wand 1997-2009 Loading required package: flowStats Loading required package: fda Loading required package: splines Loading required package: zoo Loading required package: mvoutlier Loading required package: cluster* I tried two analyze two different types of files: LMD and FCS. All files are ok, since I could analyze them in FlowJo, however, I cannot analyze them with iFlow. Can anybody, please, explain to me what am I doing wrong? Thank you very much! Regards, Anna -- Anna Dvorkin, Ph.D. Department of Biochemistry and Biomedical Sciences McMaster University 1200 Main Street West Hamilton, ON L8N Canada E-mail: dvorkin@mcmaster.ca [[alternative HTML version deleted]]
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Josef Spidlen ▴ 140
@josef-spidlen-3720
Last seen 9.6 years ago
Hi Anna, have you tried simply reading the file in R with flowCore? library(flowCore) x <- read.FCS('path to your LMS file') Do you get an error message or can you read it? As far as I know, an LMD file is just a list mode FCS file; we have some of these generated from Coulter's Cytomics FC500. These files sometimes cause problems in different software since they are unusual in terms of commonly containing more than 1 data set within the file. This is legal in FCS but I have also seen some missing offset information in these files in the FCS header segment, which is not FCS compliant (however, the appropriate information can sometimes still be derived from info in the text segment). TreeStar guys put a lot of effort into reading basically all the files out there so if a file is successfully read in FlowJo does not really tell you whether the file is OK or not. If you want, you can send me one of the files offline and I can have a closer look at it. Cheers, Josef > ------------------------------ > > Message: 2 > Date: Fri, 6 Aug 2010 11:50:15 -0400 > From: Anna Dvorkin<dvorkin at="" mcmaster.ca=""> > To: bioconductor at stat.math.ethz.ch > Subject: [BioC] iFlow error in opening FCS and LMD files > Message-ID: > <aanlktik1k40ksbf5p6xdqbq1ffvgl3qa-ljmm8_ws1f8 at="" mail.gmail.com=""> > Content-Type: text/plain > > Dear All, > > I have been trying to analyze files I obtained from a flow cytometer. In > order to do that I am trying to use the iFlow package. > > > After I write the following: > *library(iFlow)* > > A GUI window opens and I choose the files I want to analyse. > However, after I click on "Open" in the GUI, I receive the following > message: > > *"Error: '.SigArgs' is shorter than '.SigLength' says it should be"* > After I close that window, I can see, that the following packages were > successfully loaded: > > *Loading required package: RGtk2 > Loading required package: flowViz > Loading required package: lattice > KernSmooth 2.23 loaded > Copyright M. P. Wand 1997-2009 > Loading required package: flowStats > Loading required package: fda > Loading required package: splines > Loading required package: zoo > Loading required package: mvoutlier > Loading required package: cluster* > > I tried two analyze two different types of files: LMD and FCS. All files are > ok, since I could analyze > them in FlowJo, however, I cannot analyze them with iFlow. > > Can anybody, please, explain to me what am I doing wrong? > > Thank you very much! > Regards, > Anna > > > > -- > Anna Dvorkin, Ph.D. > Department of Biochemistry and Biomedical Sciences > McMaster University > 1200 Main Street West > Hamilton, ON L8N > Canada > E-mail: dvorkin at mcmaster.ca > > [[alternative HTML version deleted]] > > -- Josef Spidlen, Ph.D. Terry Fox Laboratory, BC Cancer Agency 675 West 10th Avenue, V5Z 1L3 Vancouver, BC, Canada Tel: +1 (604) 675-8000 x 7755 http://www.terryfoxlab.ca/people/rbrinkman/josef.aspx
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Hello Josef, Thank you very much for the information! Following your advice, I tried to use the flowCore package, and I was able to load one of the files I have. It was loaded without any problems. Following is the output: library(flowCore) x <- read.FCS('path to LMD file') flowFrame object 'A0-5 015.LMD' with 35117 cells and 6 observables: name desc range minRange maxRange $P1 FS LIN FS LIN 1024 0.000000 1023 $P2 SS LIN SS LIN 1024 0.000000 1023 $P3 FL3 LIN PI LIN 1024 0.000000 1023 $P4 AUX PI PEAK 1024 0.000000 1023 $P5 FL3 LOG PI LOG 1024 1.000922 10000 $P6 TIME TIME 1024 0.000000 1023 139 keywords are stored in the 'descripton' slot > summary(x) FS LIN SS LIN FL3 LIN AUX FL3 LOG TIME Min. 4.0 1.0 25.0 23 244.9 0.00 1st Qu. 233.0 145.0 392.0 343 3851.0 22.00 Median 333.0 222.0 495.0 404 4866.0 52.00 Mean 402.6 284.8 584.1 449 5725.0 56.81 3rd Qu. 509.0 352.0 795.0 531 7772.0 89.00 Max. 1023.0 1023.0 1023.0 1023 10000.0 134.00 However, when I tried to visualize the data with the flowViz package, after typing: library(flowViz) plot(x) I get the following error: Error in x@parameters$name : '.SigArgs' is shorter than '.SigLength' says it should be Actually, I am interested in analyzing cell cycle data, and I thought that I would be able to do that with the iFlow or flowCore + flowViz packages. Is there another option that can be helpful for that? Thank you in advance! Regards, Anna [[alternative HTML version deleted]]
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Hi Anna, I just tried one of our LMD files and it seems fine with both, flowCore and flowViz. So I would hope some of the flowViz guys could provide more insight in terms of what could the problem be. Generally, I think it should be possible to use flow* Bioconductor packages for cell cycle analysis. Alternatively, there's commercial software that could be used. Sorry, I have no experience there. I think some people mentioned that they used tools from verity software house. The paid version of Cyflogic should also be able to that. Cheers, Josef On 10-08-17 11:21 AM, Anna Dvorkin wrote: > Hello Josef, > Thank you very much for the information! > Following your advice, I tried to use the flowCore package, and I was > able to load one of the files I have. It was loaded without any > problems. Following is the output: > library(flowCore) > x <- read.FCS('path to LMD file') > flowFrame object 'A0-5 015.LMD' > with 35117 cells and 6 observables: > name desc range minRange maxRange > $P1 FS LIN FS LIN 1024 0.000000 1023 > $P2 SS LIN SS LIN 1024 0.000000 1023 > $P3 FL3 LIN PI LIN 1024 0.000000 1023 > $P4 AUX PI PEAK 1024 0.000000 1023 > $P5 FL3 LOG PI LOG 1024 1.000922 10000 > $P6 TIME TIME 1024 0.000000 1023 > 139 keywords are stored in the 'descripton' slot > > summary(x) > FS LIN SS LIN FL3 LIN AUX FL3 LOG TIME > Min. 4.0 1.0 25.0 23 244.9 0.00 > 1st Qu. 233.0 145.0 392.0 343 3851.0 22.00 > Median 333.0 222.0 495.0 404 4866.0 52.00 > Mean 402.6 284.8 584.1 449 5725.0 56.81 > 3rd Qu. 509.0 352.0 795.0 531 7772.0 89.00 > Max. 1023.0 1023.0 1023.0 1023 10000.0 134.00 > > However, when I tried to visualize the data with the flowViz package, > after typing: > library(flowViz) > plot(x) > I get the following error: > Error in x@parameters$name <mailto:x@parameters$name> : > '.SigArgs' is shorter than '.SigLength' says it should be > > Actually, I am interested in analyzing cell cycle data, and I thought > that I would be able to do that with the iFlow or flowCore + flowViz > packages. > Is there another option that can be helpful for that? > Thank you in advance! > Regards, > Anna -- Josef Spidlen, Ph.D. Terry Fox Laboratory, BC Cancer Agency 675 West 10th Avenue, V5Z 1L3 Vancouver, BC, Canada Tel: +1 (604) 675-8000 x 7755 http://www.terryfoxlab.ca/people/rbrinkman/josef.aspx [[alternative HTML version deleted]]
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Dear Anna [sorry that this comment is only on form, not substance, but:] Can you - provide the output of "sessionInfo()" (to see whether your package versions match and are sufficiently up-to-date) - demonstrate your problem on data files that are publicly visible (either by making some of yours so, by putting them on the web somewhere, or by finding public ones that produce the same error). I think this would greatly help the people here to see what your problem might be. Best wishes Wolfgang Wolfgang Huber EMBL http://www.embl.de/research/units/genome_biology/huber On Aug/17/10 8:38 PM, Josef Spidlen wrote: > Hi Anna, > I just tried one of our LMD files and it seems fine with both, flowCore > and flowViz. So I would hope some of the flowViz guys could provide more > insight in terms of what could the problem be. Generally, I think it > should be possible to use flow* Bioconductor packages for cell cycle > analysis. Alternatively, there's commercial software that could be used. > Sorry, I have no experience there. I think some people mentioned that > they used tools from verity software house. The paid version of Cyflogic > should also be able to that. > > Cheers, > Josef > > On 10-08-17 11:21 AM, Anna Dvorkin wrote: >> Hello Josef, >> Thank you very much for the information! >> Following your advice, I tried to use the flowCore package, and I was >> able to load one of the files I have. It was loaded without any >> problems. Following is the output: >> library(flowCore) >> x<- read.FCS('path to LMD file') >> flowFrame object 'A0-5 015.LMD' >> with 35117 cells and 6 observables: >> name desc range minRange maxRange >> $P1 FS LIN FS LIN 1024 0.000000 1023 >> $P2 SS LIN SS LIN 1024 0.000000 1023 >> $P3 FL3 LIN PI LIN 1024 0.000000 1023 >> $P4 AUX PI PEAK 1024 0.000000 1023 >> $P5 FL3 LOG PI LOG 1024 1.000922 10000 >> $P6 TIME TIME 1024 0.000000 1023 >> 139 keywords are stored in the 'descripton' slot >>> summary(x) >> FS LIN SS LIN FL3 LIN AUX FL3 LOG TIME >> Min. 4.0 1.0 25.0 23 244.9 0.00 >> 1st Qu. 233.0 145.0 392.0 343 3851.0 22.00 >> Median 333.0 222.0 495.0 404 4866.0 52.00 >> Mean 402.6 284.8 584.1 449 5725.0 56.81 >> 3rd Qu. 509.0 352.0 795.0 531 7772.0 89.00 >> Max. 1023.0 1023.0 1023.0 1023 10000.0 134.00 >> >> However, when I tried to visualize the data with the flowViz package, >> after typing: >> library(flowViz) >> plot(x) >> I get the following error: >> Error in x at parameters$name<mailto:x at="" parameters$name=""> : >> '.SigArgs' is shorter than '.SigLength' says it should be >> >> Actually, I am interested in analyzing cell cycle data, and I thought >> that I would be able to do that with the iFlow or flowCore + flowViz >> packages. >> Is there another option that can be helpful for that? >> Thank you in advance! >> Regards, >> Anna > >
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Dear Wolfgang, The packages I have are as following: > sessionInfo() R version 2.11.1 (2010-05-31) i386-pc-mingw32 locale: [1] LC_COLLATE=English_United States.1252 [2] LC_CTYPE=English_United States.1252 [3] LC_MONETARY=English_United States.1252 [4] LC_NUMERIC=C [5] LC_TIME=English_United States.1252 attached base packages: [1] splines stats graphics grDevices utils datasets methods [8] base other attached packages: [1] iFlow_2.2.0 flowStats_1.4.0 cluster_1.12.3 mvoutlier_1.4 [5] fda_2.2.2 zoo_1.6-4 flowViz_1.10.0 lattice_0.18-3 [9] flowCore_1.12.0 rrcov_1.0-00 pcaPP_1.7 mvtnorm_0.9-8 [13] robustbase_0.5-0-1 Biobase_2.6.1 RGtk2_2.12.18 loaded via a namespace (and not attached): [1] cairoDevice_2.13 feature_1.2.4 graph_1.26.0 [4] grid_2.11.1 KernSmooth_2.23-3 ks_1.6.13 [7] latticeExtra_0.6-11 MASS_7.3-6 RColorBrewer_1.0-2 [10] stats4_2.11.1 tools_2.11.1 Unfortunately, I do not have an option to post files on the web. This the first time I'm using the bioconductor mailing list, therefore I am wondering - is it possible to attach a file to the email I'm sending to the mailing list? Thank you very much! Regards, Anna [[alternative HTML version deleted]]
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