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al, International Journal of Technology and Engineering Science [IJTES]TM

Volume 2[6], pp: 1947-1951, June 2014

Geometrical Analysis of Leukocyte Nucleus to Detect

Lymphoblast from Microscopic Blood Images

Nagabhushana R M, 2Rajeshwari P

M.Tech in Bio-medical Signal Processing and Instrumentation, SJCE, Mysore, Karnataka, India
Assistant professor department of instrumentation, SJCE, Mysore, Karnataka, India.

AbstractIn order to improve patient diagnosis

various image processing software are developed to
extract useful information from medical images.
Hematologist makes the microscopic study of human
blood which led to a need of methods, including
microscope color imaging, segmentation, classification
that can allow the identification of patients suffering
from leukemia. Leukemia is related with blast white
blood cell (WBC). The nonspecific nature of the signs
and symptoms of acute lymphoblastic leukemia( ALL)
often leads to wrong diagnosis so hematologist also
find difficulty for classification hence manual
classification of blood cells is time-consuming and
susceptible to error. Therefore fast, accurate and
automatic identification of different blood cells is
required. This paper has proposed automatic Otsus
threshold used to segment the nucleus part of a white
blood cell along with image enhancement and
arithmetic operations for WBC nucleus segmentation
then the features of segmented nucleus is extracted to
classify. SVM classifier has been utilized to classify the
Index TermsLeukemia, Acute Lymphoblastic
Leukemia (ALL), White Blood Cells (WBC), Support
Vector Machine (SVM), Segmentation, Otsus method,
Morphological Features.
Medical imaging refers to the techniques and
processes used to create images of the human body for
clinical purposes such as medical procedures for
diagnosing or examining disease and also for medical
science which include the study of normal anatomy and
physiology [8]. The importance of medical analysis and
visualization can be recognized from the recent works
in the medical imaging technologies like segmentation
of kidney from ultrasound images, segmentation of
brain images [6], locating of tumors and other
pathologies [5]. Leukemia is a disease that affects blood
forming cells in the body. Its cancerous condition is
characterized by an abundance of abnormal white blood
cells in the body. Based on the research by M. D.
Anderson Cancer Center, leukemia begins in the bone
marrow and spreads to the other parts of the body [4].
In a healthy body, bone marrow makes white blood

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cells to help the body fight an infection. When a person

has leukemia, the bone marrow starts to build a lot of
abnormal white blood cells called leukemia cells. They
grow faster than normal cells and they do not stop
growing when they should.
In 2009, it is estimated that approximately 31,490
individuals will be diagnosed with leukemia and 44,510
individuals will die of the disease in the United States.
In Malaysia, a total of 21,773 cancer cases were
diagnosed among Malaysians in Peninsular Malaysia in
2006 and registered in the National Cancer Registry.
Out of this value, 3.6% of male and 1.9% of female
were diagnosed with leukemia.
The segmentation and classification of blood cells
from microscopic images allow Evaluation and
diagnosis of many diseases. Leukemia is a blood cancer
that can be detected through the analysis of WBCs or
leukocytes. Leukemia can be of Two types: acute and
chronic. According to the French-American-British
(FAB) Classification model, acute leukemia is
classified into two subtypes: acute lymphoblastic
leukemia (ALL) and acute myeloid leukemia (AML). In
this paper we consider only the ALL, The ALL
primarily affects children and adults over 50 years and
due to its rapid expansion into the bloodstream and vital
organs can be fatal if left untreated [12]. Therefore, it
becomes crucial early diagnosis of the disease for
patients' recover, especially in the case of children. The
use of image processing techniques can help to provide
information on the cells morphology. These techniques
require only one image and are therefore less
expensive, but at the same time more scrupulous in
providing more accurate standards. The main goal of
this work is the processing and analysis of microscopic
images, in order to provide a fully automatic procedure
to support the medical activity and classify.
In pathology manual detection of leukemia is done
which is time consuming as well as costly due to high
cost pathology instruments. Hence automatic technique
is adopted for fast and accurate results. In this technique
image of blood sample is processed and nucleus part is
segmented and finally cells are classified whether they
are blast or normal. The segmentation step is very
crucial because the accuracy of the subsequent feature
extraction and classification depends on the correct


Nagabhushana.R.M.et.al, International Journal of Technology and Engineering Science [IJTES]TM

Volume 2[6], pp: 1947-1951, June 2014
segmentation of white blood cells. It is also a difficult
and challenging problem due to the complex nature of
the cells and uncertainty in the microscopic image.
Many researchers have given different methods for
image segmentation. Cseke used automatic thresholding
method. Threshold techniques cannot always produce
meaningful results since no spatial information is used
during the selection of the segmentation threshold [13].
Edge detection method can also be meaningful for
segmentation but it is applicable only when there is
good contrast between foreground and background.
(Piuri and Scotti) [9]. The K-Mean clustering method is
utilized by Sinha and Ramakrishna. However, the
method of cropping the entire cell in order to get the
real area of the whole cell is not clearly shown [10].
Theera-Umpon used a fuzzy C-Mean clustering to
segment single cell images of white blood cells in the
bone marrow into two regions, i.e., nucleus and nonnucleus. The computational time increases if the
clusters number is greater than 2 [7]. In this work we
used thresholding along with morphological operators
to segment and SVM classifier is used for better
performance of classification.
A segmentation method: figure (1) shows the block
diagram of a proposed system it consists of various
functional modules. The main two steps in the proposed
system are image segmentation and classification. The
input image of blood slide is fed to the system. Blood
has four main elements to ensure it fulfills its functions
they are red blood cells, white blood cells, platelets,
plasma [1]. White blood cells fall into five categories:
Neutrophil, Eosinophil, Basophil, Monocyte and
Lymphocyte [2].White blood cell composition and
concentration in the blood gives valuable information
and plays a crucial role in the diagnosis of different
diseases. Hence the very first module in the proposed
system is image acquisition and followed by leukocyte
nucleus segmentation, then features of lymphocyte are
extracted from the segmented binary image the features
extracted are used for classification as normal or blast.
Figure(2) shows the block diagram of the
nucleus segmentation of leukocytes or WBCs. The
WBCs segmentation was made by converting the color
microscopic image to grayscale image, to enhance the
contrast of a grayscale image we used histogram
equalization which is given by the equation (1)

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Microscopic blood sub image

Leukocyte nucleus segmentation

Feature extraction
Figure 1. Block diagram of Proposed Work


( n j / n)

j 0

Where sk is the resultant histogram equalized

output and nj is the pixel value of jth pixel and n is the
total number of pixel. and the next step is contrast
stretching is a simplest piecewise linear function
contrast stretching is used to enhance the dynamic
range of a image because the images are having poor
illumination so to remove the noise of illumination
contrast stretching is used to the original microscopic
sub images .after contrast stretching to enhance the
contrast of a leukocyte we used some arithmetic
operations like addition and subtraction of both
histogram equalized image as well as contrast stretched
image To reduce noise, preserve edges and increase the
darkness of the nuclei implement 3-by-3 minimum
filter on the resultant image. The segmentation is
realized using a threshold where white regions are
clearly shows the leukocyte nucleus and black as the
There are many threshold techniques available
in literature [11] here we used Otsus thresholding [14]
A measure of region homogeneity is variance (i.e.,
regions with high homogeneity will have low variance).
Otsus method selects the threshold by minimizing the
within-class variance of the two groups of pixels
separated by the thresholding operator. It does not
depend on modeling the probability density functions,
however, it assumes a bimodal distribution of graylevel values (i.e., if the image approximately fits this
constraint, it will do a good job).Otsu threshold is
calculated by minimizes the weighted within-class
variance. This is given by equation (2)


Nagabhushana.R.M.et.al, International Journal of Technology and Engineering Science [IJTES]TM

Volume 2[6], pp: 1947-1951, June 2014
and back ground of a grayscale image. Once the
threshold value is calculated then depending upon the
value grayscale image is converted in to binary image.
The binary image have some unwanted regions we have
to remove these regions we used some morphological
operations .Edge detection is used to determine the
edges of a segmented image canny edge detection is
used because of its accuracy in detection of edges, once
the edges are determined then next step is to do image
opening which is given by the equation (9)
Where A is the image to be opened by the structuring
element B, opening is nothing but erosion followed by
dilution. Finally by using filling of connected
components by holes done to fill the connected
component to get a complete segmented image .

B Feature Extraction

Figure 2.Block Diagram Leukocyte Segmentation


(t ) q1 (t )


(t ) q2 (t )


(t )

q1 (t )

p(i )

i 1

q2 (t )

p(i )

i t 1

(t )
i 1


(t )
i t

ip(i )
q1 (t )

ip(i )
1 2 (t )


(t )


(t )]2

i 1

p(i )
q1 (t )


(t )


i t 1

(t )]2

q2 (t )

Feature extraction means to transfer the input data into

different set of features. In image processing this is a
technique of redefining a large set of redundant data
into a set of features (or feature vector) of reduced
dimension .In this paper four features of lymphocyte
cells have been observed Shape Features According to
hematologist the shape of the nucleus is an essential
feature for discrimination of blasts. Region and
boundary based shape features are extracted for shape
analysis of the nucleus. All the features are extracted
from the binary equivalent image of the nucleus with
none zero pixels representing the nucleus region. The
quantitative evaluation of each nucleus is done using
the extracted features under two classes i.e. region
based and boundary based. The features are as follows:
AREA: The area was determined by counting the total
number of none zero pixels within the image region.
PERIMETER: It was measured by calculating distance
between successive boundary pixels.
COMPACTNESS: Compactness or roundedness is the
measure of a nucleus as given by
Compactness =Perimeter2/area
SOLIDITY: The ratio of actual area and convex hull
area is known as solidity and is also an essential feature
for blast cell classification given by
Solidity =area/ convex area
ECCENTRICITY: This parameter is used to measure

where q1(t) and q2(t) in equation (3) and (4) are the
class probabilities of fore ground and background
respectively.1(t) , 2(t) in equation (5) and (6) are the
mean of fore ground and background 21(t), 22(t) in
equation (7) and (8 )are the variance of the fore ground

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how much a shape of a nucleus deviates from being

circular. Its an important feature since lymphocytes are
more circular than the blast. Eccentricity given by



Nagabhushana.R.M.et.al, International Journal of Technology and Engineering Science [IJTES]TM

Volume 2[6], pp: 1947-1951, June 2014
Where a is the major axis and b is the minor axis of the
equivalent ellipse representing the nucleus region.
ELONGATION: Abnormal bulging of the nucleus is

also a symbol which signifies leukemia. Hence the

nucleus bulging is measured in terms of a ratio called
elongation. This is defined as the ratio between
maximum distance Rmax and minimum distance Rmin
from the center of gravity to the nucleus boundary and
is given by
Elongation= Rmax/ Rmin

Where Rmaxand Rmin are maximum and minima radii

FORM FACTOR: It is a dimensionless parameter which
changes with surface irregularities and it is given by

form factor=4 pi area/ perimeter2

C. Classification
Based on the features extracted in above step,
classifier classifies the lymphocyte cells as blast or
normal cells. Classification is the task of assigning to
the unknown test vector, a label from one of the known
classes. Since the patterns are very close in the feature
space, support vector machine (SVM) support vector
machine is employed for classification. SVM is a
powerful tool for data classification based on hyper
plane classifier. This classification is achieved by a
separating surface (linear or non linear) in the input
space of the data set. They are basically two class
classifiers that optimize the margin between the classes
.The classifier training algorithm is a procedure to find
the support vectors. SVM has been utilized to classify
blast cells from normal white blood cells.

Automated systems based on artificial vision methods
can speed up this operation and they can increase the
accuracy of the response also in telemedicine
applications. Unfortunately, there are not available
public image datasets to test and compare such
algorithms. In this project public dataset of blood
samples is used, specifically designed for the evaluation
and the comparison of algorithms for segmentation and
classification [10]. For each image in the dataset, the
classification of cell is given, and it is provided a
specific set of figures of merits to be processed in order
to fairly compare different algorithms when working
with the proposed dataset. The annotation of ALLIDB1 is as follows[3]. The ALLIDB1 image files are

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named with the notation ImXXX Y.jpg where XXX is a

3-digit integer counter and Y is a Boolean digit equal to
0 is no blast cells are present, and equal to 1 if at least
one blast cell is present in the image. Please note that
all images labeled with Y=0 are from for healthy
individuals, and all images labeled with Y=1 are from
ALL patients we used sub images of main image where
the sub image consisting of one leukocyte per slide.
The proposed technique has been applied on
peripheral blood smear sub images obtained from the
public dataset as mentioned earlier. A microscopic
blood sub image is considered for evaluation [10]. As
mentioned in section 3, algorithm applied to input
image. The resulting images of segmentation are shown
in figure (3).
In figure three (a1)and (a2) are input colored image
,(b1) and (b2) are the histogram equalized images,
similarly(c1)and (c2) are thesholded output images
,(d1) and (d2) are the final segmented images.









Figure (3): Experiment results: (a) original image; (b) preprocessed

image; (c) Threshold image; (c) final segmented image.


Nagabhushana.R.M.et.al, International Journal of Technology and Engineering Science [IJTES]TM

Volume 2[6], pp: 1947-1951, June 2014
Table I
The performance of the proposed CAD method


Image A

Image B

Area in pixels



Perimeter in pixels















The above table shows the features extracted from both

Images a and b where A is the lymphoblast image and
image B is the normal lymphocyte
A WBC nucleus segmentation of stained blood
smear images followed by relevant Feature extraction
for leukemia detection is the main theme of the paper.
The paper mostly concentrates on measuring area,
orientation, eccentricity, circularity, perimeter etc.
features for better detection accuracy. Leukemia
detection with the proposed features were classified
with SVM classifier. The system is applied on 144 sub
images from public dataset giving accuracy of 88%
Furthermore the system should be robust to excessive
staining cells. Results obtained encourage future works
like stain independent blood smear image segmentation
and leukemia type classification.

We would like to express our thanks to Fabio Scotti for
providing dataset.
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Electrical and Computer Engineering, University

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