Falcon 3 overdose in EC mode

Forums

Hi there,

I am really sorry to open another topic about F3 overdose in EC mode but the other one got no reply and I am really keen on getting FindDQE (v1.06) working.

Let me first say that I had absolute no problem in measuring the DQE in linear mode. For EC mode, I have measured a total dose of 40 e/A2 both in EPU and Falcon Image Reference Manager. The dose measurements I got from the two tools match perfectly. However, when I run FindDQE the DQE goes well above 1.0.

Could it be the counts multiplier? How do I calculate a correct one?

I would be happy to share the micrographs (wetransfer or drive) if that can help!

Total dose= 40 e/A2 (total: 31.613 e/px; dose 0.6 e/px/s, exposure 52.3 s)
Apix= 0.889

FindDQE log:

################################################

FindDQE - Determine DQE of a detector
V1.06 - 06.07.15

Copyright 2013 Howard Hughes Medical Institute.
All rights reserved.
Use is subject to Janelia Farm Research Campus Software Copyright 1.1
license terms ( http://license.janelia.org/license/jfrc_copyright_1_1.html )

You can input either the counts/electron
(gain conversion factor) or the total electron dose/pixel.
Enter 1 to input counts/electron, 2 for total dose/pixel
2
2

Enter total electron dose/pixel
31.613
31.61300

Enter counts multiplier (usually 1).
1
1.000000

Enter number of images
(usually 1; 2 if also giving flat field image)
2
2

Input image with pointer silhouette
Falcon3_EC_40eA_120K_fullBeamStop_1.mrc
Falcon3_EC_40eA_120K_fullBeamStop_1.mrc

Opening MRC/CCP4 file for READ...
File : Falcon3_EC_40eA_120K_fullBeamStop_1.mrc
NX, NY, NZ: 4096 4096 1
MODE : real
Min, max : 0.000000 16287.00
Mean, RMS : 3234.504 707.4712
TITLE 1: EMAN 10/02/2017 10:49

2nd (flat field) input image
Falcon3_EC_40eA_120K_noBeamStop_1.mrc
Falcon3_EC_40eA_120K_noBeamStop_1.mrc

Output residual image
out4
out4

Reading in image

DMIN,DMAX,DMEAN= 0.000000 16287.00 3234.504

Opening MRC/CCP4 file for READ...
File : Falcon3_EC_40eA_120K_noBeamStop_1.mrc
NX, NY, NZ: 4096 4096 1
MODE : real
Min, max : 2052.000 15148.00
Mean, RMS : 3373.796 278.7028
TITLE 1: EMAN 10/02/2017 10:49

DMIN,DMAX,DMEAN= 2052.000 15148.00 3373.796

Determining image threshold

Threshold = 1738.311
N, DMEAN, DRMS inside pointer = 699447 106.0155 93.16451
N, DMEAN, DRMS outside pointer = 16077769 3370.606 278.5605
Gain conversion factor (counts/e) = 106.6209

2nd image:
N, DMEAN, DRMS outside pointer = 16077769 3373.812 278.7415
Gain conversion factor (counts/e) = 106.7223

Thresholding image
Removing single pixel outliers

Number of pixel outliers = 8

Generating smooth-edged mask for NPS calculation
Tapering edges of image
Applying mask to background
Calculating noise power spectrum
Calculating rotational average
Fitting noise spectrum with splines

Number of knots in spline = 100
Chi2 = 157975.6

Determining MTF from edge blurring using 5-Gaussian model

Coeff-1 Width-1 Coeff-2 Width-2 Coeff-3 Width-3 Coeff-4 Width-4 Coeff-5 Width-5 NormRMS
0.20000 10.00000 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07262
0.20000 9.00000 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07255
0.20000 1.00000 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07185
0.20000 0.54996 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07184
0.20000 0.00040 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07183
1.20000 0.00040 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07165
1.10534 0.00040 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07164
1.02585 0.00040 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07164
0.97432 0.00040 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07163
0.93488 0.00040 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07163
0.90674 0.00040 0.20000 8.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07163
0.90674 0.00040 0.20000 7.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07162
0.90674 0.00040 0.20000 3.00000 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07161
0.90674 0.00040 0.20000 5.17341 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07160
0.90674 0.00040 0.20000 4.84288 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07160
0.90674 0.00040 0.20000 4.45107 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07160
0.90674 0.00040 0.20000 4.32883 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07160
0.90674 0.00040 1.20000 4.32883 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07159
0.90674 0.00040 0.81746 4.32883 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07158
0.90674 0.00040 0.74727 4.32883 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07158
0.90674 0.00040 0.67891 4.32883 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07158
0.90674 0.00040 0.65699 4.32883 0.20000 6.00000 0.20000 4.00000 0.20000 2.00000 0.07158
0.90674 0.00040 0.65699 4.32883 0.20000 5.00000 0.20000 4.00000 0.20000 2.00000 0.07157
0.90674 0.00040 0.65699 4.32883 0.20000 3.34844 0.20000 4.00000 0.20000 2.00000 0.07156
0.90674 0.00040 0.65699 4.32883 0.20000 3.14270 0.20000 4.00000 0.20000 2.00000 0.07156
0.90674 0.00040 0.65699 4.32883 0.20000 3.03745 0.20000 4.00000 0.20000 2.00000 0.07156
0.90674 0.00040 0.65699 4.32883 1.20000 3.03745 0.20000 4.00000 0.20000 2.00000 0.07154
0.90674 0.00040 0.65699 4.32883 2.20000 3.03745 0.20000 4.00000 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.80675 3.03745 0.20000 4.00000 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.20000 4.00000 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.20000 3.00000 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.20000 1.54167 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 1.20000 1.54167 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.93678 1.54167 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.84882 1.54167 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 0.20000 2.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 0.20000 3.00000 0.07153
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 0.20000 2.61009 0.07152
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 1.20000 2.59103 0.07152
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 2.20000 2.59103 0.07152
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 2.90870 2.59103 0.07152
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 4.13882 2.59103 0.07152
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 5.24065 2.59103 0.07152
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 6.53865 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 7.94335 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 9.61511 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 11.40053 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 13.37934 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 14.61150 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 15.84497 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 25.84497 2.59103 0.07151
0.90674 0.00040 0.65699 4.32883 1.99692 3.03745 0.80844 1.54167 21.21415 2.59103 0.07151
0.90513 0.02321 0.65595 4.33721 1.99282 3.04421 0.80705 1.52902 21.16620 2.58968 0.07151
0.90121 0.07873 0.65341 4.35759 1.98285 3.06065 0.80367 1.49825 21.04952 2.58640 0.07151
0.90026 0.09212 0.65280 4.36250 1.98044 3.06462 0.80286 1.49084 21.02140 2.58561 0.07151
0.89675 0.14174 0.65053 4.38072 1.97152 3.07932 0.79984 1.46333 20.91710 2.58268 0.07151
1.54128 0.14727 0.65028 4.38276 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.03142 0.14727 0.65028 4.38276 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 0.65028 4.38276 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 0.65028 2.26283 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 1.65028 2.26283 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 1.90116 2.26283 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 11.90116 2.26283 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 7.31833 2.26283 1.97053 3.08096 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.79950 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.23539 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.13296 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.02086 1.46027 20.90548 2.58235 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.02086 1.46027 20.90548 2.55785 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.02086 1.46027 21.90548 2.56460 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.02086 1.46027 31.90548 2.56460 0.07151
2.18437 0.14727 7.31833 2.26283 2.97053 3.12706 0.02086 1.46027 28.57940 2.56460 0.07151
2.21801 0.32858 7.34008 2.08813 3.05604 2.98608 0.00809 1.72398 29.57940 2.59273 0.07151
2.20130 0.09224 7.32928 2.17490 3.01357 3.05610 0.00629 1.59300 29.08272 2.57876 0.07151
1.95836 0.09224 7.32928 2.17490 3.01357 3.05610 0.00629 1.59300 29.08272 2.57876 0.07151
1.97873 0.09224 7.32928 2.12827 3.01357 3.05610 0.00629 1.59300 29.08272 2.57876 0.07151
1.97873 0.09224 8.32928 2.12827 3.01357 3.05610 0.00629 1.59300 29.08272 2.57876 0.07151
1.97873 0.09224 10.12272 2.12827 3.01357 3.05610 0.00629 1.59300 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.01357 2.84679 0.00629 1.59300 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 4.01357 2.84679 0.00629 1.59300 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 0.99371 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 1.99371 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 11.99371 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 21.99371 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 31.99371 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 41.99371 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 51.99371 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 61.99372 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 66.99372 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 76.99372 2.40439 29.08272 2.57876 0.07151
1.97873 0.09224 10.00980 2.12827 3.63431 2.84679 76.99372 2.40439 31.08272 2.59611 0.07151
0.26963 0.09224 10.00980 2.12827 3.63431 2.84679 76.99373 2.40439 31.08273 2.59611 0.07151
0.90842 0.09224 10.00980 2.12827 3.63431 2.84679 76.99373 2.40439 31.08273 2.59611 0.07151
0.99756 0.09224 13.82143 2.14891 3.63431 2.63545 76.99373 2.40439 31.08273 2.59611 0.07151
0.99756 0.09224 13.82143 2.14891 7.62181 2.63545 92.99373 2.40439 31.08273 2.59611 0.07151
0.01639 0.09224 17.63306 2.16956 11.60930 2.42410 108.99371 2.40439 31.77512 2.59611 0.07151
0.10838 0.09224 17.27572 2.24590 11.23548 2.44391 107.49371 2.40439 31.74267 2.59611 0.07151

Final values:

Coeff-1 Width-1 Coeff-2 Width-2 Coeff-3 Width-3 Coeff-4 Width-4 Coeff-5 Width-5 NormRMS
0.10838 1.50776 20.14814 2.24171 27.23548 2.44392 107.49371 2.40439 31.74267 2.59611 0.07151

Opening MRC/CCP4 file for WRITE...
File : out4
NX, NY, NZ: 1242 1242 1
MODE : real

RMS of noise adjusted for the MTF = 562.2076

DQE(0) = 1.138072

Frequency MTF NPS NPS-Fit DQE
0.000000 1.000000 2.000000 1.000000 1.138072
0.000488 1.000000 0.417012 0.922861 1.233198
0.000977 0.999998 2.000000 0.861337 1.321281
0.001465 0.999996 0.868825 0.813885 1.398309
0.001953 0.999993 0.931630 0.778964 1.460986
0.002441 0.999989 0.658171 0.755031 1.507284
0.002930 0.999984 1.074720 0.740546 1.536752
0.003418 0.999978 0.570644 0.733965 1.550512
0.003906 0.999971 0.605467 0.733749 1.550949
0.004395 0.999964 0.896250 0.738353 1.541252
0.004883 0.999955 0.775330 0.746238 1.524942
0.005371 0.999946 0.682556 0.755860 1.505501
0.005859 0.999935 0.714238 0.765679 1.486164
0.006348 0.999924 0.916084 0.774152 1.469866
0.006836 0.999912 0.729182 0.779858 1.459075
0.007324 0.999899 0.469100 0.782536 1.454043
0.007813 0.999885 0.916090 0.782566 1.453948
0.008301 0.999870 0.739374 0.780335 1.458061
0.008789 0.999855 0.759422 0.776231 1.465724
0.009277 0.999838 0.928942 0.770641 1.476307
0.009766 0.999820 0.854221 0.763952 1.489181
0.010254 0.999802 0.885564 0.756552 1.503692
0.010742 0.999783 0.721879 0.748828 1.519143
0.011230 0.999763 0.670484 0.741168 1.534783
0.011719 0.999741 0.696758 0.733958 1.549793
0.012207 0.999719 0.862361 0.727587 1.563296
0.012695 0.999696 0.739810 0.722441 1.574359
0.013184 0.999673 0.664263 0.718900 1.582038
0.013672 0.999648 0.679760 0.717092 1.585949
0.014160 0.999622 0.643467 0.716845 1.586413
0.014648 0.999596 0.754168 0.717973 1.583837
0.015137 0.999569 0.642012 0.720288 1.578659
0.015625 0.999540 0.680094 0.723603 1.571339
0.016113 0.999511 0.663494 0.727730 1.562337
0.016602 0.999481 0.834520 0.732481 1.552109
0.017090 0.999450 0.783330 0.737670 1.541096
0.017578 0.999418 0.821208 0.743108 1.529721
0.018066 0.999386 0.791995 0.748609 1.518381
0.018555 0.999352 0.748925 0.753984 1.507455
0.019043 0.999317 0.692310 0.759046 1.497298
0.019531 0.999282 0.762689 0.763609 1.488247
0.020020 0.999246 0.734119 0.767537 1.480522
0.020508 0.999208 0.738588 0.770842 1.474064
0.020996 0.999170 0.700915 0.773560 1.468774
0.021484 0.999131 0.875447 0.775724 1.464560
0.021973 0.999091 0.761770 0.777372 1.461340
0.022461 0.999050 0.831697 0.778536 1.459034
0.022949 0.999009 0.827030 0.779254 1.457569
0.023438 0.998966 0.753556 0.779559 1.456874
0.023926 0.998922 0.816995 0.779488 1.456880
0.024414 0.998878 0.742107 0.779075 1.457523
0.024902 0.998833 0.755823 0.778355 1.458739
0.025391 0.998787 0.718783 0.777363 1.460464
0.025879 0.998739 0.763800 0.776136 1.462636
0.026367 0.998692 0.779106 0.774707 1.465193
0.026855 0.998643 0.826605 0.773119 1.468059
0.027344 0.998593 0.731993 0.771413 1.471159
0.027832 0.998542 0.744287 0.769632 1.474414
0.028320 0.998491 0.743768 0.767818 1.477744
0.028809 0.998438 0.783075 0.766015 1.481068
0.029297 0.998385 0.854616 0.764264 1.484303
0.029785 0.998331 0.655327 0.762608 1.487363
0.030273 0.998276 0.811525 0.761090 1.490165
0.030762 0.998219 0.787057 0.759752 1.492621
0.031250 0.998163 0.823123 0.758638 1.494644
0.031738 0.998105 0.701377 0.757788 1.496146
0.032227 0.998046 0.768941 0.757247 1.497040
0.032715 0.997986 0.766723 0.757054 1.497242
0.033203 0.997926 0.752618 0.757211 1.496751
0.033691 0.997865 0.803139 0.757668 1.495665
0.034180 0.997802 0.794113 0.758373 1.494086
0.034668 0.997739 0.738944 0.759277 1.492119
0.035156 0.997675 0.726150 0.760327 1.489866
0.035645 0.997610 0.736725 0.761472 1.487432
0.036133 0.997544 0.746230 0.762661 1.484917
0.036621 0.997477 0.767122 0.763842 1.482423
0.037109 0.997410 0.683845 0.764964 1.480048
0.037598 0.997341 0.770289 0.765976 1.477889
0.038086 0.997272 0.756285 0.766827 1.476045
0.038574 0.997202 0.722560 0.767464 1.474612
0.039063 0.997130 0.815494 0.767837 1.473686
0.039551 0.997058 0.822579 0.767912 1.473327
0.040039 0.996985 0.792110 0.767725 1.473470
0.040527 0.996911 0.742017 0.767325 1.474020
0.041016 0.996837 0.771005 0.766761 1.474882
0.041504 0.996761 0.739985 0.766084 1.475962
0.041992 0.996685 0.822605 0.765343 1.477166
0.042480 0.996607 0.740360 0.764587 1.478396
0.042969 0.996529 0.808547 0.763866 1.479559
0.043457 0.996449 0.817555 0.763230 1.480556
0.043945 0.996369 0.760534 0.762728 1.481292
0.044434 0.996289 0.749927 0.762411 1.481668
0.044922 0.996207 0.693723 0.762327 1.481587
0.045410 0.996124 0.682611 0.762527 1.480953
0.045898 0.996040 0.815634 0.763057 1.479676
0.046387 0.995956 0.753014 0.763915 1.477762
0.046875 0.995870 0.781916 0.765061 1.475297
0.047363 0.995784 0.788903 0.766450 1.472367
0.047852 0.995697 0.730203 0.768041 1.469059
0.048340 0.995609 0.771277 0.769791 1.465460
0.048828 0.995520 0.780904 0.771658 1.461655
0.049316 0.995430 0.815735 0.773597 1.457727
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0.995605 0.149617 0.055765 0.053939 0.472313
0.996094 0.149332 0.056115 0.053794 0.471784
0.996582 0.149048 0.055816 0.053649 0.471261
0.997070 0.148764 0.055978 0.053504 0.470743
0.997559 0.148481 0.056518 0.053358 0.470231
0.998047 0.148198 0.054593 0.053212 0.469723
0.998535 0.147915 0.056918 0.053066 0.469220
0.999023 0.147633 0.054696 0.052920 0.468721
0.999512 0.147351 0.054375 0.052774 0.468226
1.000000 0.147070 0.054503 0.052628 0.467735

################################################

For a counting detector, you could determine the pixel variance and compare it with the exposure per pixel that you think you have. The average variance should be close to the average pixel value (Poisson distribution). If this is not true, it would point to some kind of multiplier.

In reply to by niko

Hi,
I also am getting values for DQE on our Falcon3 EC which start close to 2. There is a multiplier which FEI uses so images scale to 16 bit integers, and seems to be about 100.

Here are the stats for a 100s image at a dose rate of 0.85 e/pix/sec: (85e/pix)
e2iminfo.py -s 18may03e_00030enn.mrc
18may03e_00030enn.mrc 1 images in MRC format 4096 x 4096
0.
apix=1.11 min=6775 max=1.167e+04 mean=9069 sigma=458.6 skewness=0.07348 kurtosis=0.005946

Variance would be 210,313.9, far different from the expected value

For comparison, here are values for an image taken on the K2 at 4 e/pix/sec, 20s exposure:
e2iminfo.py -s 18may05a_00024enn.mrc
18may05a_00024enn.mrc 1 images in MRC format 3710 x 3838
0.
apix=0.67 min=0 max=360.4 mean=69.53 sigma=9.185 skewness=0.2264 kurtosis=0.2285
Variance would be 96, close to the expected value of 80.

Thanks for any information on this.

In reply to by wrice

Thanks for this analysis. If I understand your numbers correctly, the variance for the K2 image should be the square of sigma = 9.185, i.e. 84.4, which is indeed pretty close to 80. For the Falcon3, the variance of 210,313.9 is not close to the expected value of 85. If one assumes a multiplication of the values read out from the Falcon3 by a factor of 50, the variance would be reduced by a factor of 2500 and the corrected variance would be 84.1, close to the expected value of 85. If this is correct, users should answer 50 to the prompt "Enter counts multiplier (usually 1)". FindDQE should then correct for this additional multiplier and (hopefully) produce the correct result.

In reply to by niko

Thanks for the fast response. I seem to get the same results even with the multiplier:
---
Final values:

Coeff-1 Width-1 Coeff-2 Width-2 Coeff-3 Width-3 Coeff-4 Width-4 Coeff-5 Width-5 NormRMS
175.18721 2.16472 36.98002 2.11803 16.82003 2.05402 35.90425 1.96127 78.11725 2.05722 0.04628

Opening MRC/CCP4 file for WRITE...
File : residuals/18may03e_00031enn-res.mrc
NX, NY, NZ: 1232 1232 1
MODE : real

RMS of noise adjusted for the MTF = 1110.419

DQE(0) = 0.7849888

Frequency MTF NPS NPS-Fit DQE
0.000000 1.000000 2.000000 1.000000 0.784989
0.000488 1.000000 1.191514 0.909434 0.863161
0.000977 0.999999 2.000000 0.828804 0.947132
0.001465 0.999997 0.433388 0.757539 1.036229
0.001953 0.999994 1.662521 0.695066 1.129360
0.002441 0.999991 0.669550 0.640813 1.224966
0.002930 0.999987 0.906638 0.594210 1.321029
0.003418 0.999982 0.377233 0.554683 1.415154
...
---

Here was my input:
--
Enter 1 to input counts/electron, 2 for total dose/pixel
2

Enter total electron dose/pixel
85.00000

Enter counts multiplier (usually 1).
50.00000

Enter number of images
(usually 1; 2 if also giving flat field image)
2

Input image with pointer silhouette
18may03e_00031enn.mrc

Opening MRC/CCP4 file for READ...
File : 18may03e_00031enn.mrc
NX, NY, NZ: 4096 4096 1
MODE : real
Min, max : 11.00000 11943.00
Mean, RMS : 8688.720 1861.290
--

In reply to by niko

Thanks Niko, I still get similar results:

----

You can input either the counts/electron
(gain conversion factor) or the total electron dose/pixel.
Enter 1 to input counts/electron, 2 for total dose/pixel
2

Enter total electron dose/pixel
85.00000

Enter counts multiplier (usually 1).
2.0000000E-02

Enter number of images
(usually 1; 2 if also giving flat field image)
2

Input image with pointer silhouette
18may03e_00031enn.mrc

Opening MRC/CCP4 file for READ...
File : 18may03e_00031enn.mrc
NX, NY, NZ: 4096 4096 1
MODE : real
Min, max : 11.00000 11943.00
Mean, RMS : 8688.720 1861.290

2nd (flat field) input image
18may03e_00030enn.mrc

Output residual image
residuals/18may03e_00031enn-res.mrc

Reading in image

DMIN,DMAX,DMEAN= 11.00000 11943.00 8688.720
File : 18may03e_00030enn.mrc
NX, NY, NZ: 4096 4096 1
MODE : real
Min, max : 6775.000 11673.00
Mean, RMS : 9068.908 458.5512

DMIN,DMAX,DMEAN= 6775.000 11673.00 9068.909

Determining image threshold

Threshold = 4659.799
N, DMEAN, DRMS inside pointer = 733361 244.9134 219.1284
N, DMEAN, DRMS outside pointer = 16043855 9074.685 461.0667
Gain conversion factor (counts/e) = 2.135220

2nd image:
N, DMEAN, DRMS outside pointer = 16043855 9066.184 458.5136
Gain conversion factor (counts/e) = 2.133220

Thresholding image
Removing single pixel outliers

Number of pixel outliers = 10

...
RMS of noise adjusted for the MTF = 1110.405

DQE(0) = 0.7850079

Frequency MTF NPS NPS-Fit DQE
0.000000 1.000000 2.000000 1.000000 0.785008
0.000488 1.000000 1.191463 0.909437 0.863179
0.000977 0.999998 2.000000 0.828810 0.947148
0.001465 0.999997 0.433436 0.757546 1.036244
0.001953 0.999994 1.662540 0.695075 1.129373
0.002441 0.999991 0.669573 0.640823 1.224977
0.002930 0.999987 0.906647 0.594220 1.321038
0.003418 0.999982 0.377215 0.554693 1.415161
0.003906 0.999977 0.649167 0.521671 1.504726
0.004395 0.999970 0.627984 0.494580 1.587126
0.004883 0.999964 0.374117 0.472851 1.660039
0.005371 0.999956 0.538788 0.455910 1.721699
0.005859 0.999948 0.380352 0.443185 1.771100

In reply to by wrice

Thanks for posting the data. The numbers in the images suggest that they represent the counts multiplied by about 50. If I assume an exposure of 170 e/pixel, i.e. twice the exposure you indicated, I obtain the following DQE curve and residuals image:

Falcon 3 DQE curve at 300 kV

Falcon 3 DQE residuals at 300 kV

I understand that the exposure was provided by the EPU. Is there a way to establish it independently of the microscope software and hardware? It looks to me as if the displayed exposures given by the microscope are a factor of 2 too low.

In reply to by niko

Thanks! I was using the Gain Reference Manager program to estimate dose, so perhaps something must be miscalibrated there. Kinds of disturbing if we are dosing with twice what we think we are. I could collect a long exposure under the same conditions on the K2. I suppose if I correct for the magnification difference caused by the different placements in the column I could calculate back.

In reply to by wrice

Dear Niko,

could you please upload also the output image to the dqe curve that you calculated using double the dose. I would like to see how this image should look like if the calculation succeeded correctly. I also have problems measuring the dqe using find dqe and different dose rates.

In reply to by cwigge

Another thing that does not work is when I provide 2 images. find_dqe complains that the dose is different. When I run e2iminfo.py -s I indeed get different mean values for images taken on empty parts of the grid with and without beamstop using TIA.

output is:

e2iminfo.py -s 0.3_beamstop.tif 0.3_empty.tif
0.3_beamstop.tif 1 images in TIFF format 4096 x 4096
0.
apix=1.00 min=0 max=6.528e+04 mean=5.06e+04 sigma=1.103e+04 skewness=-3.631 kurtosis=13.85

0.3_empty.tif 1 images in TIFF format 4096 x 4096
0.
apix=1.00 min=0 max=6.528e+04 mean=3.099e+04 sigma=1.118e+04 skewness=0.1234 kurtosis=-0.0777

2 total images
representing 0 particles
cwigge@ubuntu:~/Desktop/DQE_TEST$ e2iminfo.py -s 0.3_beamstop.mrc 0.3_empty.mrc
0.3_beamstop.mrc 1 images in MRC format 4096 x 4096
0.
apix=1.00 min=0 max=6.528e+04 mean=5.06e+04 sigma=1.103e+04 skewness=-3.631 kurtosis=13.85

0.3_empty.mrc 1 images in MRC format 4096 x 4096
0.
apix=1.00 min=0 max=6.528e+04 mean=3.099e+04 sigma=1.118e+04 skewness=0.1234 kurtosis=-0.0777

2 total images
representing 0 particles
cwigge@ubuntu:~/Desktop/DQE_TEST$

Do you know why this is the case? Could that affect the measurement? How can I solve this?

In reply to by cwigge

I posted the residuals image.

The different means presumably reflect that fact that the pointer image has an area with low counts while the other image does not. The error message produced by FindDQE should reflect differences in areas of the two images that do not contain the pointer. The means in these areas should not exceed 5%.