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This commit is contained in:
moonanjum26
2018-08-10 12:05:28 +05:30
parent 188e46804b
commit 126057ce23
8 changed files with 0 additions and 721 deletions
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Flowcytometer_Focusing_Jeevan2018/flowcytometergui.fig
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Flowcytometer_Focusing_Jeevan2018/flowcytometergui.m
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function varargout = flowcytometergui(varargin)
% FLOWCYTOMETERGUI MATLAB code for flowcytometergui.fig
% FLOWCYTOMETERGUI, by itself, creates a new FLOWCYTOMETERGUI or raises the existing
% singleton*.
%
% H = FLOWCYTOMETERGUI returns the handle to a new FLOWCYTOMETERGUI or the handle to
% the existing singleton*.
%
% FLOWCYTOMETERGUI('CALLBACK',hObject,eventData,handles,...) calls the local
% function named CALLBACK in FLOWCYTOMETERGUI.M with the given input arguments.
%
% FLOWCYTOMETERGUI('Property','Value',...) creates a new FLOWCYTOMETERGUI or raises the
% existing singleton*. Starting from the left, property value pairs are
% applied to the GUI before flowcytometergui_OpeningFcn gets called. An
% unrecognized property name or invalid value makes property application
% stop. All inputs are passed to flowcytometergui_OpeningFcn via varargin.
%
% *See GUI Options on GUIDE's Tools menu. Choose "GUI allows only one
% instance to run (singleton)".
%
% See also: GUIDE, GUIDATA, GUIHANDLES
% Edit the above text to modify the response to help flowcytometergui
% Last Modified by GUIDE v2.5 09-Jul-2018 11:27:06
% Begin initialization code - DO NOT EDIT
gui_Singleton = 1;
gui_State = struct('gui_Name', mfilename, ...
'gui_Singleton', gui_Singleton, ...
'gui_OpeningFcn', @flowcytometergui_OpeningFcn, ...
'gui_OutputFcn', @flowcytometergui_OutputFcn, ...
'gui_LayoutFcn', [] , ...
'gui_Callback', []);
if nargin && ischar(varargin{1})
gui_State.gui_Callback = str2func(varargin{1});
end
if nargout
[varargout{1:nargout}] = gui_mainfcn(gui_State, varargin{:});
else
gui_mainfcn(gui_State, varargin{:});
end
% End initialization code - DO NOT EDIT
% --- Executes just before flowcytometergui is made visible.
function flowcytometergui_OpeningFcn(hObject, eventdata, handles, varargin)
% This function has no output args, see OutputFcn.
% hObject handle to figure
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
% varargin command line arguments to flowcytometergui (see VARARGIN)
% Choose default command line output for flowcytometergui
handles.output = hObject;
s = serial('COM12');
set(s,'BaudRate',57600); set(s,'Terminator','#');
set(s,'Timeout',30); % s.RecordName = 'CytoSerialTxnLog.txt';
fopen(s); % record(s);
handles.s = s;
msg=strcat(datestr(clock,'yyyy-mm-dd-HHMM'),'m',datestr(clock,'ss'),'s');
fid = fopen(strcat(msg,'.txt'),'w');
handles.fid = fid;
if (fid == -1)
disp('could not open logging file. You probably need to change working directory to the one where the MATLAB file is located.');
return;
end
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% Start cam
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
tic
fprintf(fid, '\r\nStarting camera...');
% choose which webcam (winvideo-1) and which mode (YUY2_176x144)
%vid = videoinput('winvideo', 1, 'RGB32_744x480');
vid = videoinput('winvideo', 2, 'RGB24_640x480');
% Configure the object for manual trigger mode.
triggerconfig(vid, 'manual');
% only capture one frame per trigger, we are not recording a video
vid.FramesPerTrigger = 1;
% output would image in RGB color space
vid.ReturnedColorspace = 'rgb';
% tell matlab to start the webcam on user request, not automatically
triggerconfig(vid, 'manual');
% we need this to know the image height and width
vidRes = get(vid, 'VideoResolution');
% image width
imWidth = vidRes(1);
% image height
imHeight = vidRes(2);
% number of bands of our image (should be 3 because it's RGB)
nBands = get(vid, 'NumberOfBands');
% create an empty image container and show it on axPreview
hImage = image(zeros(imHeight, imWidth, nBands), 'parent', handles.axPreview);
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% do auto white balance
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
src = getselectedsource(vid);
% src.BalanceWhiteAuto = 'Continuous';
% begin the webcam preview
currzpos = 0;
preview(vid, hImage);
% expose the objects so we can handle them in other functions
handles.vid = vid;
handles.src = src;
handles.currz = currzpos;
guidata(hObject,handles);
%start(vid);
drawnow;
% % set and display the exposure
% src.ExposureTime = 360;
%set(handles.expDisp,'string', src.ExposureTime );
Time_to_init_cam = toc
% Update handles structure
guidata(hObject, handles);
% UIWAIT makes flowcytometergui wait for user response (see UIRESUME)
% uiwait(handles.figure1)
% --- Outputs from this function are returned to the command line.
function varargout = flowcytometergui_OutputFcn(hObject, eventdata, handles)
% varargout cell array for returning output args (see VARARGOUT);
% hObject handle to figure
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
% Get default command line output from handles structure
varargout{1} = handles.output;
% --- Executes on button press in autofocus.
function autofocus_Callback(hObject, eventdata, handles)
% hObject handle to autofocus (see GCBO)
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
s = handles.s; vid = handles.vid; currzpos = handles.currz;
[~, ~, ~] = focusstack_1(s,vid,currzpos);pause(2)
% --- Executes on button press in moveleft.
function moveleft_Callback(hObject, eventdata, handles)
% hObject handle to moveleft (see GCBO)
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
s = handles.s; fprintf(s,'A')
% --- Executes on button press in moveright.
function moveright_Callback(hObject, eventdata, handles)
% hObject handle to moveright (see GCBO)
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
s = handles.s; fprintf(s,'D')
% --- Executes on button press in snapshot.
function snapshot_Callback(hObject, eventdata, handles)
% hObject handle to snapshot (see GCBO)
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
vid = handles.vid;
snapshot = getsnapshot(vid);
imwrite(snapshot,'snap.jpg')
function figure1_DeleteFcn(hObject, eventdata, handles)
% hObject handle to figure1 (see GCBO)
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
if ~isempty(instrfind)
fclose(instrfind);
delete(instrfind);
end
try
fid=handles.fid; fclose(fid);
vid = handles.vid; stoppreview(vid); delete(vid);
catch
clear all;
clc;
end
%s=handles.s; fclose(s); delete(s);
disp('Killed all before exiting');
% --- Executes on button press in startvid.
function startvid_Callback(hObject, eventdata, handles)
% hObject handle to startvid (see GCBO)
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
vid = handles.vid;
vid_trigger = 1; handles.vid_trigger = vid_trigger;
for x=1:1000
snap = getsnapshot(vid)
imwrite(strcat('Image_',num2str(x),'.jpg'),snap)
end
% aviObject = VideoWriter('newfile.avi','Motion JPEG AVI'); handles.aviObject = aviObject;
% open(aviObject);
% while(vid_trigger == 1)
% snap = getsnapshot(vid);
% writeVideo(aviObject,snap);
% end
% --- Executes on button press in stopvid.
function stopvid_Callback(hObject, eventdata, handles)
% hObject handle to stopvid (see GCBO)
% eventdata reserved - to be defined in a future version of MATLAB
% handles structure with handles and user data (see GUIDATA)
aviObject = handles.aviObject;
handles.vid_trigger = 0;
close(aviObject);

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Flowcytometer_Focusing_Jeevan2018/focusplane_4.m
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clear;clc;
disp('Scan Inializing........')
tic;
t1 = toc;
s = serial('COM3');
set(s,'BaudRate',57600); set(s,'Terminator','#');
set(s,'Timeout',30); %s.RecordName = 'CytoSerialTxnLog.txt';
fopen(s); %recweblistord(s);
msg=strcat(datestr(clock,'yyyy-mm-dd-HHMM'),'m',datestr(clock,'ss'),'s');
fid = fopen(strcat(msg,'.txt'),'w');
if (fid == -1)
disp('could not open logging file. You probably need to change working directory to the one where the MATLAB file is located.');
return;
end
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% Start cam
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
tic
disp('\r\nStarting camera...')
fprintf(fid, '\r\nStarting camera...');
% choose which webcam (winvideo-1) and which mode (YUY2_176x144)
%vid = videoinput('winvideo', 1, 'RGB32_744x480');
vid = videoinput('gentl', 1, 'BGRA8Packed');
% Configure the object for manual trigger mode.
triggerconfig(vid, 'manual');
% only capture one frame per trigger, we are not recording a video
vid.FramesPerTrigger = 1;
% output would image in RGB color space
vid.ReturnedColorspace = 'rgb';
% tell matlab to start the webcam on user request, not automatically
triggerconfig(vid, 'manual');
% we need this to know the image height and width
vidRes = get(vid, 'VideoResolution');
% image width
imWidth = vidRes(1);
% image height
imHeight = vidRes(2);
% number of bands of our image (should be 3 because it's RGB)
nBands = get(vid, 'NumberOfBands');
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% do auto white balance
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
src = getselectedsource(vid);
src.BalanceWhiteAuto = 'Continuous';
src.ExposureTime = 800;
start(vid);
Time_to_init_cam = toc
% this is the slide name
setslidename = 'Slide';
fprintf(s,'J');currxpos = 0;
fprintf(s,'K');currypos = 0;
fprintf(s,'L');currzpos = 0;
pause(0.1);
disp('Origin is set');
xFoV = round(1280*5.3/(34*0.159));yFoV = round(1024*5.3/(34*0.159));
xFoV_mm = 0.1995;yFoV_mm = 0.1596;
XPulseRate = 38400; YPulseRate = 38400 ;
Xzadjustdelay = 0.002;
autoContrTime = 0.1;
XImDelay = (xFoV / XPulseRate) ;
YImDelay = (yFoV / YPulseRate) ;
%at y = 12.5mm line
Ypos = strcat('+',sprintf('%06d',abs(41359)));
fprintf(s,'N'); fprintf(s,Ypos);pause(41359/YPulseRate);currypos=41359;
param_y0 =[];Zval=[];
xpos1 = 0; xpos2 = 200000;
[currzpos, temp_var] = focusstack_1(s,vid,currzpos);pause(2)
tf1 = toc;
for xpos = xpos1:17986:xpos2
Xpos = strcat('+',sprintf('%06d',abs(xpos )));
fprintf(s,'B'); fprintf(s,Xpos);pause(17986/XPulseRate)
[currzpos, temp_var] = focusstack_2(s,vid,currzpos);
Zval = [Zval currzpos];
snapshot=getsnapshot(vid);
disp('saving best focussed snapshot..')
imwrite(snapshot,strcat('images/finding_best_region/t7/t7_x_',Xpos,'_',sprintf('%06d',abs(currzpos)),'.bmp'));
param_y0 = [param_y0 parameter(snapshot)]; currxpos = xpos;
end
disp(toc-tf1)
xpos = xpos1:17986:xpos2;
[max_param_y0, Index] = max(param_y0);
xbest1 = abs(xpos(Index));
Xbest1 = strcat('+',sprintf('%06d',xbest1));
fprintf(s,'B'); fprintf(s,Xpos);pause(abs(currxpos-xbest1)/XPulseRate)
Zfoc1 = Zval(Index);gotoZ(s,Zfoc1,currzpos);currzpos=Zfoc1;param_y1 = []; Zval=[];
tf2 =toc;
if(xbest1 <= 17986)
for xpos = (xbest1):7194:(xbest1+17986*2)
Xpos = strcat('+',sprintf('%06d',abs(xpos )));
fprintf(s,'B'); fprintf(s,Xpos);pause(17986/XPulseRate)
[currzpos, temp_var] = focusstack_2(s,vid,currzpos);
Zval = [Zval currzpos];
snapshot=getsnapshot(vid);
disp('saving best focussed snapshot..')
imwrite(snapshot,strcat('images/finding_best_region/t7/t7_x_',Xpos,'_',sprintf('%06d',abs(currzpos)),'.bmp'));
param_y1 = [param_y1 parameter(snapshot)]; currxpos = xpos;
end
elseif(xbest1>=200000)
for xpos = (xbest1-17986*2):7194:(xbest1)
Xpos = strcat('+',sprintf('%06d',abs(xpos )));
fprintf(s,'B'); fprintf(s,Xpos);pause(17986/XPulseRate)
[currzpos, temp_var] = focusstack_2(s,vid,currzpos);
Zval = [Zval currzpos];
snapshot=getsnapshot(vid);
disp('saving best focussed snapshot..')
imwrite(snapshot,strcat('images/finding_best_region/t7/t7_x_',Xpos,'_',sprintf('%06d',abs(currzpos)),'.bmp'));
param_y1 = [param_y1 parameter(snapshot)]; currxpos = xpos;
end
else
for xpos = (xbest1-17986):7194:(xbest1+17986)
Xpos = strcat('+',sprintf('%06d',abs(xpos )));
fprintf(s,'B'); fprintf(s,Xpos);pause(17986/XPulseRate)
[currzpos, temp_var] = focusstack_2(s,vid,currzpos);
Zval = [Zval currzpos];
snapshot=getsnapshot(vid);
disp('saving best focussed snapshot..')
imwrite(snapshot,strcat('images/finding_best_region/t7/t7_x_',Xpos,'_',sprintf('%06d',abs(currzpos)),'.bmp'));
param_y1 = [param_y1 parameter(snapshot)]; currxpos = xpos;
end
end
disp(toc-tf2)
xpos = (xbest1-17896):7194:(xbest1+17896);
param_y1 = (param_y1>0.0999);disp(param_y1)
target1 = sum(param_y1(1:3)); target2 = sum(param_y1(3:5));
fprintf(s,'N'); fprintf(s,'0000000');pause(abs(41359)/YPulseRate);
tf3=toc;
if((sum(param_y1(2:4))==3))
xbest2 = xpos(2);xbest1 = xpos(4);
Xbest2 = strcat('+',sprintf('%06d',xbest2));
Xbest1 = strcat('+',sprintf('%06d',xbest1));
fprintf(s,'B'); fprintf(s,Xbest1);pause((abs(currxpos-xbest1))/XPulseRate);gotoZ(s,Zval(4),currzpos);currzpos=Zval(4);fprintf(s,'R');
fprintf(s,'B'); fprintf(s,Xbest2);pause((abs(17986))/XPulseRate);gotoZ(s,Zval(2),currzpos);currzpos=Zval(2);fprintf(s,'E');currxpos = xbest2;
fprintf(s,'N'); fprintf(s,'0078719');pause(abs(78719)/YPulseRate);currypos = 78719;
[currzpos, temp_var] = focusstack_2(s,vid,currzpos);fprintf(s,'Y');
fprintf(s,'Z');
xA = xbest2; xB = xbest1;
yC = 78719; yB =0;
numofX = (xB - xA) / xFoV;
numofY = (yC - yB) / yFoV;
while fscanf(s) ~= 'S'
end;
% NOW TAKING PICTURES
rowcount = 0;
filecount = 0;
msg = '\r\n Starting image capture ...';
fprintf(fid, msg);
% Dircn = false; %We believe we start from top
while(rowcount <= numofY)
while(filecount <= numofX)
%pause(XImDelay*2);
snapshot=getsnapshot(vid);
%enh_snapshot=autocontrast(snapshot);
filename = strcat(setslidename,'_',num2str(rowcount),'-',num2str(filecount), '.bmp');
%imwrite(enh_snapshot,filename);
imwrite(snapshot,strcat('images/',filename));
filecount = filecount + 1;
end
% wait for shift by one FoV in Y direction
filecount = 0;
%pause(YImDelay*2);
rowcount = rowcount + 1;
end
elseif(target1>target2)
xbest2 = xpos(1);xbest1 = xpos(3);
Xbest2 = strcat('+',sprintf('%06d',xbest2));
Xbest1 = strcat('+',sprintf('%06d',xbest1));
fprintf(s,'B'); fprintf(s,Xbest1);pause((abs(currxpos-xbest1))/XPulseRate);gotoZ(s,Zval(3),currzpos);currzpos=Zval(3);fprintf(s,'R');
fprintf(s,'B'); fprintf(s,Xbest2);pause((abs(17986))/XPulseRate);gotoZ(s,Zval(1),currzpos);currzpos=Zval(1);fprintf(s,'E');currxpos = xbest2;
fprintf(s,'N'); fprintf(s,'0078719');pause(abs(78719)/YPulseRate);currypos = 78719;
[currzpos, temp_var] = focusstack_2(s,vid,currzpos);fprintf(s,'Y');
fprintf(s,'Z');
xA = xbest2; xB = xbest1;
yC = 78719; yB =0;
numofX = (xB - xA) / xFoV;
numofY = (yC - yB) / yFoV;
while fscanf(s) ~= 'S'
end;
% NOW TAKING PICTURES
rowcount = 0;
filecount = 0;
msg = '\r\n Starting image capture ...';
fprintf(fid, msg);
% Dircn = false; %We believe we start from top
while(rowcount <= numofY)
while(filecount <= numofX)
%pause(XImDelay*2);
snapshot=getsnapshot(vid);
%enh_snapshot=autocontrast(snapshot);
filename = strcat(setslidename,'_',num2str(rowcount),'-',num2str(filecount), '.bmp');
%imwrite(enh_snapshot,filename);
imwrite(snapshot,strcat('images/',filename));
filecount = filecount + 1;
end
% wait for shift by one FoV in Y direction
filecount = 0;
%pause(YImDelay*2);
rowcount = rowcount + 1;
end
elseif(target1<target2)
xbest2 = xpos(5);xbest1 = xpos(3);
Xbest2 = strcat('+',sprintf('%06d',xbest2));
Xbest1 = strcat('+',sprintf('%06d',xbest1));
fprintf(s,'B'); fprintf(s,Xbest1);pause((abs(currxpos-xbest1))/XPulseRate);gotoZ(s,Zval(3),currzpos);currzpos=Zval(3);fprintf(s,'E');
fprintf(s,'B'); fprintf(s,Xbest2);pause((abs(17986))/XPulseRate);gotoZ(s,Zval(5),currzpos);currzpos=Zval(5);fprintf(s,'R');currxpos = xbest2;
fprintf(s,'N'); fprintf(s,'0078719');pause(abs(78719)/YPulseRate);currypos = 78719;
[currzpos, temp_var] = focusstack_2(s,vid,currzpos);fprintf(s,'Y');
fprintf(s,'Z');
xA = xbest1; xB = xbest2;
yC = 78719; yB =0;
numofX = (xB - xA) / xFoV;
numofY = (yC - yB) / yFoV;
while fscanf(s) ~= 'S'
end;
% NOW TAKING PICTURES
rowcount = 0;
filecount = 0;
msg = '\r\n Starting image capture ...';
fprintf(fid, msg);
% Dircn = false; %We believe we start from top
while(rowcount <= numofY)
while(filecount <= numofX)
%pause(XImDelay*2);
snapshot=getsnapshot(vid);
%enh_snapshot=autocontrast(snapshot);
filename = strcat(setslidename,'_',num2str(rowcount),'-',num2str(filecount), '.bmp');
%imwrite(enh_snapshot,filename);
imwrite(snapshot,strcat('images/t7/',filename));
filecount = filecount + 1;
end
% wait for shift by one FoV in Y direction
filecount = 0;
%pause(YImDelay*2);
rowcount = rowcount + 1;
end
else
disp('Slide is not prepared well!!! Please make a good slide..!')
end
disp(toc-tf3)
stop(vid);
fprintf(s,'N'); fprintf(s,'0000000');
fprintf(s,'B'); fprintf(s,'0000000');
fprintf(s,'M'); fprintf(s,'0000000');
pause(0.1)
if ~isempty(instrfind)
fclose(instrfind);
delete(instrfind);
end
t2 = toc;
disp((t2-t1)/60)

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Flowcytometer_Focusing_Jeevan2018/focusstack_1.m
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function [Zval, Variance, currzpos] = focusstack_1(s,vid,z)
t1 = toc;currzpos=z;
snap = getsnapshot(vid);
imwrite(snap,strcat('images/withouthc/snap_at_start','.bmp'))
%fprintf(s,'L');
% CRUDE AF.
Z_travel_for_crude = 16000; crude_step_count = 200;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
crude_pulse_count = Z_travel_for_crude / crude_step_count;
crude_max_var=0; crude_loc_max_var=0;
crude_start_loc = z-(Z_travel_for_crude/2);
% gotoZ(s,crude_start_loc,currzpos);pause(1) ;
fprintf(s,'M');pause(3)
% for i=1: crude_pulse_count/2
% fprintf(s,'T');pause(0.35)
% end
currzpos=crude_start_loc;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% now keep going up and taking var
crude_curr_loc = crude_start_loc;
Variance = []; Zval = [];
for i=1: crude_pulse_count
% variance
snap = getsnapshot(vid);
imwrite(snap,strcat('images/snap_at_',num2str(crude_curr_loc),'.bmp'))
crude_img = double(snap); crude_curr_var=var(crude_img(:));
%display(strcat('Variance',num2str(crude_curr_var),'Iter:',num2str(i)))
%pause(0.5)
Variance = [Variance crude_curr_var];
Zval = [Zval crude_curr_loc];
if (crude_curr_var > crude_max_var)
crude_max_var = crude_curr_var;
crude_loc_max_var = crude_curr_loc;
end
% update loc
crude_curr_loc = crude_curr_loc + crude_step_count ;
fprintf(s,'G');pause(0.1)
%gotoZ(s,crude_curr_loc,currzpos);
currzpos=crude_curr_loc;
end
disp(currzpos)
disp(crude_loc_max_var)
crude_pulse_count = round(abs(crude_loc_max_var - currzpos)/200);
for i=1: crude_pulse_count
if(currzpos>crude_loc_max_var)
fprintf(s,'T');pause(0.1)
currzpos = currzpos - 200;
elseif(currzpos<crude_loc_max_var)
fprintf(s,'G');pause(0.1)
currzpos = currzpos + 200;
end
end
pause(1)
snap = getsnapshot(vid);
imwrite(snap,strcat('images/withouthc/snap_at_end_crude','.bmp'))
t2 = toc;
disp(t2-t1)
end

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Flowcytometer_Focusing_Jeevan2018/focusstack_2.m
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function [Zval, Variance] = focusstack_2(s,vid,z)
t1 = toc;currzpos=z;
% fINECRUDE AF.
Z_travel_for_crude = 6000; crude_step_count = 200;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
crude_pulse_count = Z_travel_for_crude / crude_step_count;
crude_max_var=0;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% go to starting Z loc (we've hit limit switch via arduino already so its z=0)
crude_start_loc = z- (Z_travel_for_crude/2);
gotoZ(s,crude_start_loc,currzpos); currzpos=crude_start_loc;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% now keep going up and taking var
crude_curr_loc = crude_start_loc;
for i=1: crude_pulse_count
% variance
crude_img = double(getsnapshot(vid)); crude_curr_var=var(crude_img(:));
if (crude_curr_var > crude_max_var)
crude_max_var = crude_curr_var;
crude_loc_max_var = crude_curr_loc;
end
% update loc
crude_curr_loc = crude_curr_loc + crude_step_count ;
gotoZ(s,crude_curr_loc,currzpos); currzpos=crude_curr_loc;
end
gotoZ(s,crude_loc_max_var,currzpos);currzpos=crude_loc_max_var;
% fINECRUDE AF.
Z_travel_for_crude = 500; crude_step_count = 50;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
crude_pulse_count = Z_travel_for_crude / crude_step_count;
crude_max_var=0;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% go to starting Z loc (we've hit limit switch via arduino already so its z=0)
crude_start_loc = currzpos- (Z_travel_for_crude/2);
gotoZ(s,crude_start_loc,currzpos);currzpos=crude_start_loc;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% now keep going up and taking var
crude_curr_loc = crude_start_loc;
for i=1: crude_pulse_count
% variance
crude_img = double(getsnapshot(vid)); crude_curr_var=var(crude_img(:));
if (crude_curr_var > crude_max_var)
crude_max_var = crude_curr_var;
crude_loc_max_var = crude_curr_loc;
end
% update loc
crude_curr_loc = crude_curr_loc + crude_step_count ;
gotoZ(s,crude_curr_loc,currzpos);currzpos=crude_curr_loc;
end
gotoZ(s,crude_loc_max_var,currzpos);currzpos=crude_loc_max_var;
% The fine AF begins here
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
Ztotaltravel = 100; final_go_backpulses = 0 ;
currZ = crude_loc_max_var;
% first go above
start_above_loc = currZ- (Ztotaltravel/2);
gotoZ(s,start_above_loc,currzpos);currzpos=start_above_loc;
% now build the var array
localAF_loc_array = zeros(1,Ztotaltravel); localAF_var_array = zeros(1,Ztotaltravel);
localAF_curr_loc = start_above_loc;
for q=1:(Ztotaltravel/5)
% location
localAF_loc_array(q) = localAF_curr_loc;
% variance
localAF_img = double(getsnapshot(vid)); localAF_var_array(q)=var(localAF_img(:));
% move and update loc
localAF_curr_loc = localAF_curr_loc + 5; gotoZ(s,localAF_curr_loc,currzpos);
currzpos=localAF_curr_loc;
end
% Finally go to the max var location
index_of_max_var = localAF_var_array == max(localAF_var_array);
loc_of_max_var = localAF_loc_array(index_of_max_var) - final_go_backpulses ;
gotoZ(s,loc_of_max_var,currzpos); currzpos=loc_of_max_var;
Zval = loc_of_max_var;
Variance = max(localAF_var_array);
t2 = toc;
disp(t2-t1)
end

60
Flowcytometer_Focusing_Jeevan2018/focusstack_3.m
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@@ -1,60 +0,0 @@
function [Zval, Variance] = focusstack_3(s,vid,z)
t1 = toc;currzpos=z;
% fINECRUDE AF.
Z_travel_for_crude = 1000; crude_step_count = 50;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
crude_pulse_count = Z_travel_for_crude / crude_step_count;
crude_max_var=0;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% go to starting Z loc (we've hit limit switch via arduino already so its z=0)
crude_start_loc = z - (Z_travel_for_crude/2);
gotoZ(s,crude_start_loc,currzpos); currzpos=crude_start_loc;
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
% now keep going up and taking var
crude_curr_loc = crude_start_loc;
for i=1: crude_pulse_count
% variance
crude_img = double(getsnapshot(vid)); crude_curr_var=var(crude_img(:));
if (crude_curr_var > crude_max_var)
crude_max_var = crude_curr_var;
crude_loc_max_var = crude_curr_loc;
end
% update loc
crude_curr_loc = crude_curr_loc + crude_step_count ;
gotoZ(s,crude_curr_loc,currzpos); currzpos=crude_curr_loc;
end
gotoZ(s,crude_loc_max_var,currzpos);currzpos=crude_loc_max_var;
% The fine AF begins here
%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%
Ztotaltravel = 100; final_go_backpulses = 0 ;
currZ = crude_loc_max_var;
% first go above
start_above_loc = currZ - (Ztotaltravel/2);
gotoZ(s,start_above_loc,currzpos);currzpos=start_above_loc;
% now build the var array
localAF_loc_array = zeros(1,Ztotaltravel); localAF_var_array = zeros(1,Ztotaltravel);
localAF_curr_loc = start_above_loc;
for q=1:(Ztotaltravel/5)
% location
localAF_loc_array(q) = localAF_curr_loc;
% variance
localAF_img = double(getsnapshot(vid)); localAF_var_array(q)=var(localAF_img(:));
% move and update loc
localAF_curr_loc = localAF_curr_loc + 5; gotoZ(s,localAF_curr_loc,currzpos);
currzpos=localAF_curr_loc;
end
% Finally go to the max var location
index_of_max_var = localAF_var_array == max(localAF_var_array);
loc_of_max_var = localAF_loc_array(index_of_max_var) - final_go_backpulses ;
gotoZ(s,loc_of_max_var,currzpos);currzpos=loc_of_max_var;
Zval = loc_of_max_var;
Variance = max(localAF_var_array);
t2 = toc;
disp(t2-t1)
end

14
Flowcytometer_Focusing_Jeevan2018/gotoZ.m
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@@ -1,14 +0,0 @@
function gotoZ(s,check,currzpos)
ZPulseRate = 38400;
if(check<200000 && check>-200000)
str = sprintf('%06d',abs(round(check))); % abs needed so that no +&- in neg numbers
if((check) < 0)
newstr = strcat('-',str);
else
newstr = strcat('+',str);
end
disp(strcat('moving to position:',newstr))
% end formatting ---------- --------------------------------------------
fprintf(s,'M'); fprintf(s, newstr ); pause(0.035);
end
end

21
Flowcytometer_Focusing_Jeevan2018/parameter.m
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@@ -1,21 +0,0 @@
function param = parameter(image)
t1 = toc;
bw = rgb2gray(image);
[Gx, Gy] = imgradientxy(bw);
%[Gmag, Gdir] = imgradient(Gx, Gy);
gradImage = rgb2gray(imfuse(Gx,Gy));
%temp = histeq(gradImage);
threshImage = im2bw(gradImage,graythresh(gradImage));
[~, radii] = imfindcircles(threshImage,[10 60]);
cell_area = round(sum(3.14*(radii.^2)));
bg = rgb2gray(imread('bg.jpg'));
a = histeq(bw)-bg;
level2 = graythresh(a);
bgImg = bwareaopen(im2bw(a,level2),800);
bg_pixels = sum(sum(bgImg == 1));
fg_pixels = (1280 * 1024) - bg_pixels;
fg_pixels = fg_pixels - round(0.1*fg_pixels);
param = cell_area/fg_pixels;
t2 = toc;
disp(t2-t1)
end