poresize_analysis_finalimages.m

% Pore size segmentation for final data
clear;
% clc;

basedir='/Users/sayantanbhatttacharya/Library/CloudStorage/Box-Box/2021.12.15 Images for Alternate Analyses/cryoSEM/Analyzed Images Final/';
basedir2='/Users/sayantanbhatttacharya/Library/CloudStorage/Box-Box/2021.12.15 Images for Alternate Analyses/cryoSEM/';
geltype={'HAX30','C2HX30','C6HX30','C4','C4HX20','C4HX30','C4HAX40'};
% Xbar=categorical(geltype);
G=length(geltype);
trials={'Replicate 1','Replicate 2','Replicate 3'};
T=length(trials);
ycrop=880;
magnification=17;%(%pixels/microns)
meanporesize=zeros(G,T,3);
stdporesize=zeros(G,T,3);
poresize=cell(G,T,3);

excelcol={'A2','B2','C2'};

savedir=[basedir2,'Results/','segmentimage/'];
if ~exist(savedir,'dir')
    mkdir(savedir);
end

%Load images in a loop
for i=1:G
    for j=1:T

        imagedir=fullfile([basedir,geltype{i},filesep,trials{j}]);

        list_ims=dir(fullfile(imagedir,'*.tif'));
        Nim=length(list_ims);
%         writecell(trials(j),'poresize.xlsx','Sheet',geltype{i},'WriteMode','append')

        for n=1:Nim

            savename=[savedir,geltype{i},'_',trials{j},'_',list_ims(n,1).name(1:end-4)];

            img1=imread([imagedir,filesep,list_ims(n,1).name]);
            img1crop=img1(1:ycrop,:);

            if i==1
                [BW,maskedImage] = segmentImageHAX(img1crop);
            else
                [BW,maskedImage] = segmentImage(img1crop);
            end
% %             figure;imagesc(~maskedImage);
% % 
% %             img1filtered2=stdfilt(im2double(img1crop),true(5));
% %             img1filtered2(img1filtered2<0.05)=0;
% %             figure;imagesc(img1filtered2);
% %             img1filtered2=stdfilt(im2double(imadjust(img1crop)),true(5));
% %             img1filtered2(img1filtered2<0.05)=0;
% %             figure;imagesc(img1filtered2);

            [label2,meanarea,stddevarea,Numofelem,deq]=poresize_calculate(BW,magnification,img1crop,savename,geltype{i});
            pause(0.01);

            meanporesize(i,j,n)=meanarea/magnification;
            stdporesize(i,j,n)=stddevarea/magnification;
            poresize{i,j,n}=deq/magnification;

            writecell(poresize(i,j,n)','poresize2.xlsx','Sheet',geltype{i},'WriteMode','append')


        end

        writecell(trials(j),'poresize2.xlsx','Sheet',geltype{i},'WriteMode','append')

    end
end
% keyboard;
% save([savedir,'poresize_allimages.mat'],'poresize');
%%
meanporesizearray=reshape(meanporesize,G,T*3);
meanporesize1=mean(meanporesizearray,2);
meanporesize1(1)=mean(meanporesizearray(1,1:8));

stdporesizearray=reshape(stdporesize,G,T*3);
stdporesize1=mean(stdporesizearray,2);
stdporesize1(1)=mean(stdporesizearray(1,1:8));

errhigh=stdporesize1;
errlow=stdporesize1;

Xbar1=categorical({'HAX30','C2HX30','C4HX30','C6HX30'});
Xbar1=reordercats(Xbar1,{'HAX30','C2HX30','C4HX30','C6HX30'});
Xbar2=categorical({'C4','C4HX20','C4HX30','C4HAX40'});
Xbar2=reordercats(Xbar2,{'C4','C4HX20','C4HX30','C4HAX40'});
figure;
bar(Xbar1',meanporesize1([1 2 6 3]));
hold on;
er = errorbar(Xbar1',meanporesize1([1 2 6 3]),errlow([1 2 6 3]),errhigh([1 2 6 3]));    
er.Color = [0 0 0];                            
er.LineStyle = 'none';
er.LineWidth=3;
hold off;
ylabel('Pore Size (\mum)');
ylim([0 10]);
set(gca,'FontSize',20);
% print(gcf,'-dpng',[savedir,'poresie_1.png'],'-r200');


figure;
bar(Xbar2',meanporesize1([4 5 6 7]));
hold on;
er = errorbar(Xbar2',meanporesize1([4 5 6 7]),errlow([4 5 6 7]),errhigh([4 5 6 7]));    
er.Color = [0 0 0];                            
er.LineStyle = 'none';  
er.LineWidth=3;
hold off;
ylabel('Pore Size (\mum)');
ylim([0 10]);
set(gca,'FontSize',20);
% print(gcf,'-dpng',[savedir,'poresie_2.png'],'-r200');
%%
% geltype
[meanporesize1 stdporesize1]

% save([savedir,'poresize_allimages.mat'],'poresize','meanporesize1','stdporesize1');
%%
function [label2,meanarea,stddevarea,Numofelem,deq]=poresize_calculate(binary_img,magnification,img1crop,savename,geltype)
% BW=imbinarize(im2double(img1));
%% Connected components
CC = bwconncomp(binary_img,8);
% CC = bwconncomp(BW,8);
label2 = labelmatrix(CC);
stats = regionprops(CC,'Centroid','Area','MajorAxisLength','MinorAxisLength');

area1=([stats.Area]);
area1=sqrt(area1*4/pi);%Equivalent Diameter conversion

area1(area1<=0.2*magnification)=[];%remove diameters less than 0.3 microns
area1(area1>=40*magnification)=[];%remove diameters greater than 40 microns

Numofelem=length(area1);
meanarea=mean(area1(1:end));
stddevarea=std(area1(1:end));
deq=area1;

%% plotting
figure(12);
tiledlayout(1,3,'TileSpacing','tight');
% histogram(area1(2:end),0:10:250); 
nexttile;imshow(labeloverlay(img1crop,label2));
nexttile;imshow(img1crop);hold on;plot((1:10)*magnification+500,500*ones(10,1),'r-','LineWidth',3);hold off;
title(geltype);
bins1=0:1:10;
nexttile;histogram(area1/magnification,bins1); axis square;
title(['mean diameter=',num2str(meanarea/magnification,'%03.2f'),'\mum']);
% set(gcf,'position',[100,100,900,300]);
set(gcf,'position',[100,700,1200,600]);
% print(gcf,'-dpng',[savename,'_segmented_img_overlay.png']);

% img1filtered2=entropyfilt(img1crop,true(3));
% figure;imagesc(img1filtered2);

if meanarea==0
    keyboard;
end

end

% close all;
% figure(11);imshow(labeloverlay(img1,label2));
% % figure(11);imshow(img1);
% % hold on;
% % imshow(label2rgb(label2,'jet','k','shuffle'));
% % title('Connected Components Over Original Image');
% % hold off;
% print(gcf,'-dpng',[basedir,Cname,filesep,'labelmatrix_new2_seg_inv_5000_02.png']);

% BWoutline = bwperim(label2);
% Segout = img1; 
% Segout(BWoutline) = 255; 
% figure;imshow(Segout)
% title('Outlined Original Image')
% figure;imagesc(label2);colormap('hot')
% print(gcf,'-dpng',[basedir,Cname,filesep,'labelmatrix2_seg_inv_5000_02.png']);
% area1=sqrt([stats.Area]/pi);