Abstract:

Malaria is mosquito-borne blood disease caused by parasites of the genus

Plasmodium. Conventional diagnostic tool for malaria is the examination of stained
blood cell of patient in microscope. The blood to be tested is placed in a slide and is
observed under a microscope to count the number of infected RBC. An expert
technician is involved in the examination of the slide with intense visual and mental
concentration. This is tiresome and time consuming process.
In this paper, we construct a new mage processing system for detection and
quantification of plasmodium parasites in blood smear slide, later we develop
Machine Learning algorithm to learn, detect and determine the types of infected cells
according to its features

5
 Chapter 1

INTRODUCTION:

The study proposes an image processing model for detection of malaria infected cells.
We use image processing techniques to detect parasite-infected red blood cells in
thin smears on standard microscope slides The most widely used present day method
is examining thin blood smears under a microscope, and visually searching for
infected cells. A clinician manually counts the number of parasitic red blood cells -
sometimes up to 5,000 cells (according to WHO protocol).Malaria could be
forestalled, controlled, and relieved all the more adequately if an increasingly precise
and effective symptomatic techniques were accessible. We have utilized image
processing procedures to identify the nearness of malaria contaminated cells.And to
classify the stage of malaria i.e falciparum or non-falciparum we use machine
learning.
Hundreds of millions of blood films are examined every year for malaria, which
involves manual counting of parasites and infected red blood cells by a trained
microscopist. Accurate parasite counts are essential not only for malaria diagnosis.
They are also important for testing for drug resistance, measuring drug-effectiveness,
and classifying disease severity. However, microscopic diagnostics is not
standardized and depends heavily on the experience and skill of the microscopist.It is
common for micro scopists in low-resource settings to work in isolation, with no
rigorous system in place that can ensure the maintenance of their skills and thus
diagnostic quality.This leads to incorrect diagnostic decisions in the field.For false-
negative cases, this leads to unnecessary use of antibiotics, a second consultation, lost
days of work, and in some cases progression into severe malaria. For false positive
cases, a misdiagnosis entails unnecessary use of anti-malaria drugs and suffering from
their potential side effects, such as nausea, abdominal pain, diarrhea, and sometimes
severe complications.

6
 Chapter 2

LITERATURE REVIEW

2.1 Malaria Detection using Image Processing and Machine Learning

Abstract

Malaria is a deadly, infectious and lifethreatening mosquito-borne blood disease

caused by Plasmodium parasites. The conventional and most standard way of
diagnosing malaria is by visually examining blood smears via microscope for
parasite-infected red blood cells under the microscope by qualified technicians. This
method is inefficient and time consuming and the diagnosis depends on the
experience and the knowledge of the person doing the examination. Automatic image
recognition technologies based on image processing have been applied to malaria
blood smears for diagnosis before. However, the practical performance has not been
up to the mark so far. This gives us all the motivation to make malaria detection and
diagnosis fast, easy and efficient. Our main aim is to build a model that can detect
cells from images of multiple cells in thin blood smear on standard microscope slides
and classify them as either infected or uninfected with early and effective testing
using image processing. And also perform classification on the infected cell image
using machine learning.

INTRODUCTION

Malaria is a deadly, infectious disease caused by the Plasmodium parasite which is

transmitted by the bites of female Anopheles mosquitoes. According to the World
Malaria Report 2019 published by WHO, there were an estimated 405,000 malaria
related deaths in the preceding year. The disease is curable but early detection holds
the key. Existing methods used to detect Malaria include microscopic detection of
infected cells in a laboratory. The method is both expensive and tedious. An estimated
93 percent of all Malaria cases in 2018 were reported in the WHO African region. The
region also has one of the lowest per capita incomes across the world. A faster, low
cost, and reliable alternative to microscopic detection of Malaria is proposed in this
model.

Problem Statement

We propose an image processing model for detection of malaria infected cells. We

use image processing techniques to detect parasite-infected red blood cells in thin
smears on standard microscope slides. The most widely used present day method is
analyzing thin blood smears under a microscope, and visually searching for
contaminated cells. A clinician manually counts the number of parasitic red blood
cells - sometimes up to 5,000 cells (according to WHO protocol). Malaria could be
forestalled, controlled, and relieved all the more adequately if an increasingly precise
and effective symptomatic techniques were accessible. We have utilized image
processing procedures to identify the nearness of malaria contaminated cells. And to
classify the stage of malaria whether it is falciparum which is the most deadliest stage
in malaria or non-falciparum, for this we use machine learning technologies.

7
Scope of the Project

The Malaria Detection from thin film blood smear images demands segmentation of
single blood cells from the microscopic blood slide images which can be taken from a
pathologist and the dataset would contain cell images that are not segmented. Hence,
segmentation in the proposed method is done using a variety of image processing
techniques. Edge detection techniques and segmentation techniques used in this
system overcomes the issue of overlapping of cells by eliminating the noise and
finding the discontinuities of the cells. It differentiates each cell and detects the
infection in the cell using morphological segmentation. Also, all the images are raw
and have different intensities, and since there is no uniformity in all the images,
detection of cells and infection is very difficult. To overcome this problem, the
proposed method uses histogram matching where all the images are standard and has
the same intensity which in turn increases the accuracy level.

METHODOLOGY
At present, the recognition of Malaria parasite in single cell slide is totally manual.
This procedure could be rearranged by capturing an image of the blood smear and
afterwards utilizing the proposed model to arrange whether the cells are contaminated
or not. The proposed model uses the utilization of image processing systems to
improve existing techniques and abbreviate the time taken for recognition of malaria
parasite in blood tests. The dataset is manually collected from the CDC‘s Division of
parasitic infection and Malaria. Our concentration here is to make a mechanized
capacity to distinguish the nearness of Malaria parasite in slight blood spread and
measure the segment of RBC in the example that are tainted, primary assignment is to
fragment the contaminations, for which segmentation of the cells is the earlier
undertaking. Segmentation techniques involve methodologies based on Edge
Detection, Watershed Segmentation and Morphological segmentation Once the cells
are segmented, the infections are segmented. This is done via using a threshold
intensity pixel value for the infection, i.e. if the pixel value is in the range of the
threshold the infections are identified.

Segmentation of cells

When we "segment" an image, we distinguish the regions of interest (ROIs) from the
non-ROI portion, generally creating a binary mask of what we want to qualify,
quantify, track, etc. Segmentation is a critical part of many image processing
problems, and is worth considering in some depth. Here also we will segment our cell
images. One of the major issues in segmenting human blood cells is the contiguity of
the cells, one may find a lot of overlapping of cells. Segmenting touching object is
one of the most difficult task in image processing. We use different approaches like
Edge Detection, Watershed Segmentation, Morphological Segmentation.
1) Edge Detection(Gradient Based Techniques): The gradient based edge detections
look for the first derivative of an image where the maxima and minima are occur.
These techniques used sobel, prewitt and robert‘s cross operator for finding the edges.
2) Edge Detection(Gaussian Based Techniques): The main purpose of this technique
is to detect the zero crossings

8
in the second order derivative of an image to find edges. The Gaussian based
techniques are Laplacian of Gaussian (LOG) and Canny edge detection.

The LOG operator works best in all of the edge detection techniques since it can
eliminate the noise levels and discontinuities in the cells better as compared to the
other edge detection techniques which would otherwise not prove to be good for
further detection and it would be hard to segment the cells.

3) Watershed Segmentation: Watershed segmentation algorithm is applied on the

images in the dataset, it segments or divides the different cells present in the
image. These divisions help in simplifying the detection and classification of malaria
parasite in the blood samples.

4) Morphological Segmentation: The term morphology refers to the shape and size of
the objects within the image. In our study we have used the shapes of the cells, as
most of the cells were circular in shape and of a particular radii range. Thus, a circle
detection algorithm was designed which could detect the cells based on the
shape of the cells.

Segmentation of infection

After successful detection of the cells the next task is to spot the infections inside the
cells. So, for this we would be using the pixel intensity values, and observe the range
of intensities wherever the infections. But the problem with it is that the illumination
and color scale of one image may differ to other. For this we would use histogram
matching to have a uniform color scale.

Classification of infected cells

We use machine learning to classify the cells if infected. Support Vector

Machine(SVM) is a supervised machine learning algorithm that can be used for both
classification and regression tasks. An SVM classifies data by finding the best
hyperplane that separates data points of one class from those of the other class. The
best hyperplane for an SVM implies the one with the largest margin between the two
classes. Margin means the maximal width of the slab corresponding to the hyperplane
that has no interior data points.

Data Source
We used archived blood smear images acquired from the CDC‘s Division of parasitic
infection and Malaria as input. Our data set contains images of different malaria
parasites like Plasmodium Falciparum, Plasmodium Knowlesi, Plasmodium Malariae,
Plasmodium Ovale, Plasmodium Vivax.

Steps used in Image Processing

1. We apply an auto-generated segmenter that performs five steps:
(1) Convert to grayscale
(2) Initialize segmentation with Otsu‘s threshold.
(3) Filter components by area.
(4)Form masked from input image and segmented image.

9
2. Detecting edges-We have used various edge detection techniques like Canny,
Laplacian of Gaussian ,Prewitt, Roberts, Sobel and Zerocross operator out of which
LOG gives us the best result.
3. Combining the edges(logically) with the segmented regions.
4. Improve edge mask by performing morphological operations like imclose, skeleton,
etc.
5. Cleaning of the mask and refine the mask.
6. Use different segmentation techniques like watershed, thresholding, k-means
clustering based segmentation. Of which we select watershed segmentation since the
other techniques have tiny pores and do not give good results for further segmentation.
7. To improve the result and flatten the pools from the results of watershed we use
imhmin function.
8. We use another segmentation technique i.e. Morphological segmentation to
overcome the shortcomings in watershed segmentation. This technique detects the
circles by setting a specific range of radius.
9. Histogram matching is done since all the images have different intensities and for
classification we require all the images to be standard with the same intensity.
10. Finally, if the intensity values are within the threshold we display the percentage
of cells infected.

Classification Stage

We used Classification Learner to automatically train a selection of different

classification models on our data. We use automated training to quickly try a selection
of model types, then explore promising models interactively. Here, the two classes are
falciparum and non-falciparum. If the cell is infected in the image processing stage,
then we undergo a test for classifying whether the malaria parasite is falciparum or
some other malaria parasite(non-falciparum). Machine learning algorithms like Cubic
SVM, Linear SVM and cosine KNN are used and out of which the best algorithm
with the highest accuracy score is selected for classification purpose.

Outputs
All the images are read in MATLAB and an objective/target image is chosen on
which all the tasks are performed and afterward on all the images. After performing
all the steps of image processing, the images of blood cells are displayed whether
infected or not infected, and if infected the number of cells infected and their
percentage is shown and then we train the model for classification of the infected cell.

CONCLUSIONS
Considering our above methodology, we come to the conclusion that the newly
designed system of image processing technologies is suitable for parasite detection.
The morphological segmentation technique and improved watershed segmentation
technique proves to be the best in detection of the cells and segmenting them from the
nonregion of interest portion. To characterize the sort of plasmodium parasite for
which we utilized machine learning technologies and cubic SVM proves to be the best
by having the highest accuracy score. The system has been tested with around 110
thin film blood smear images and the results are satisfactory. The inconsistencies
among the images was a challenge but we have tried to make our system robust.

10
 2.2 Image analysis and machine learning for
detecting malaria

INTRODUCTION
Malaria is caused by protozoan parasites of the genus Plasmodium that are transmitted
through the bites of infected female Anopheles mosquitoes and that infect the red
blood cells. Most deaths occur among children in Africa, where a child dies almost
every minute from malaria, and where malaria is a leading cause of childhood neuro-
disability. According to the World Malaria Report 2016, an estimated 3.2 billion
people in 95 countries and territories are at risk of being infected with malaria and
developing disease, and 1.2 billion are at high risk (>1 in 1000 chance of getting
malaria in a year). There were about 214 million cases of malaria globally
in 2016 and about 438,000 malaria deaths. The burden was heaviest in the African
region, where an estimated 92% of all malaria deaths occurred, and in children aged
under 5 years, who accounted for more than two thirds of all deaths (see also the
malaria death rates from an earlier WHO report. Typical symptoms of malaria include
fever, fatigue, headaches, and, in severe cases, seizures and coma, leading to death.
Hundreds of millions of blood films are examined every year for malaria, which
involves manual counting of parasites and infected red blood cells by a trained
microscopist. Accurate parasite counts are essential not only for malaria diagnosis.
They are also important for testing for drugresistance, measuring drug-effectiveness,
and classifying disease severity. However, microscopic diagnostics is not
standardized and depends heavily on the experience and skill of the microscopist. It is
common for microscopists in low-resource settings to work in isolation, with no
rigorous system in place that can ensure the maintenance of their skills and thus
diagnostic quality. This leads to incorrect diagnostic decisions in the field. For
false-negative cases, this leads to unnecessary use of antibiotics, a second consultation,
lost days of work, and in some cases progression into severe malaria. For false
positive cases, a misdiagnosis entails unnecessary use of anti-malaria drugs and
suffering from their potential side effects, such as nausea, abdominal pain, diarrhea,
and sometimes severe complications. This sober analysis of malaria diagnosis has
prompted efforts to perform malaria diagnosis automatically. Automatic parasite
counting has several advantages compared with manual counting:
(1) it provides a more reliable and standardized interpretation of blood films,
(2) it allows more patients to be served by reducing the workload of the malaria field
workers, and
(3) (3) it can reduce diagnostic costs. Several key processing steps are typically
required to quantify parasitemia automatically. First, digital blood slide images need
to be acquired, which often requires preprocessing to normalize for lighting or
staining variations. In a second step, blood cells or parasites need to be detected. For
blood cells, this typically implies cell segmentation to identify individual cells in cell
clumps to obtain accurate cell counts. In a third step, after cell detection and
segmentation, features are computed to describe the typical visual appearance of
infected and uninfected blood cells. In a final classification step, a classifier, who has
been trained on an independent and typically manually annotated training set, then
discriminates between infected and uninfected cells. Once the number of infected and
uninfected cells is known, computation of parasitemia is a straightforward athematical
equation, which includes clinical parameters such as hematocrit value, for example.
The prospects of automating malaria diagnosis with its obvious advantages has
attracted many researchers, especially in the last decade. The publications reflect all

11
the major developments we have seen in the areas of automatic pattern recognition
and machine learning in the last years. Our article will give an overview of the articles
that have been published, using the processing steps mentioned above as a framework
and guide. This is not the first survey article on the subject. In fact, several survey
articles have already been published before, which bear testimony to both the
importance of automated malaria diagnosis and the research dynamics and rapid
system development. We refer readers in particular to the following surveys for
additional information about the background of automatic malaria diagnosis and the
image processing and machine learning methods used for automated microscopy
diagnosis of malaria.In addition, more specific surveys have been published on cell
features for malaria parasite detection, on malaria diagnosis, on malaria diagnostic
tools, and on alternatives to conventional microscopy. The purpose of our article is
not to replace these surveys, but rather to complement them and to provide the latest
update of the state of the art in image analysis and machine learning for malaria
diagnosis as it presents itself at the end of the year 2017. With about 170 literature
citations, we have collected more references compared with the other surveys. We had
the goal to include also maybe lesser known publications to provide a historical
documentation of the work done. In addition, we included a section on deep learning,
which is the latest development in malaria diagnosis and which arguably has the
potential to render many of the old approaches obsolete, similar to the development in
other imaging application areas. There have also been many developments in
hardware for automatic malaria diagnosis, which are however out of the scope of this
article and deserve a separate article.Nevertheless, we devote a section to rapid
diagnostic tests (RDTs) for malaria diagnosis because they are also widely used in
the field. The bulk of our articles have been collected from the Journal of Microscopy,
Malaria Journal, and PLOS ONE, including a few articles from Nature and others.
We have also collected publications from Institute of Electrical and Electronics
Engineers (IEEE) conferences and other proceedings published by Springer and
Elsevier. Furthermore, we have organized the articles into sections for preprocessing,
cell detection and segmentation, feature computation, and classification. We have also
added a separate section about deep learning and an ex
tensive section about mobile smartphone applications for malaria diagnosis. A
discussion of the latest developments and our conclusion mark the end of this article.

MALARIA

There are 5 Plasmodium species that cause malaria in human: Plasmodium falciparum,
Plasmodium vivax, Plasmodium malariae, Plasmodium ovale, and Plasmodium
knowlesi. The 2 most common species are P. falciparum and P. vivax. P. falciparum
is the most severe form and is responsible for most malaria-related deaths globally.1
P. falciparum is the most prevalent malaria parasite in sub-Saharan Africa, accounting
for 99% of estimated malaria cases in 2016. Outside of Africa, P. vivax is the
predominant parasite in the WHO Region of the Americas, representing 64% of
malaria cases, and is above 30% in the WHO Southeast Asia and 40% in the Eastern
Mediterranean regions.Each of these parasite species goes through stages during their
development cycle (48 hours), which gives the parasites a different visual appearance
that can be observed under the microscope. In chronologic order, these stages are the
ring stage, trophozoite stage, schizont stage, and gametocyte stage. typical examples
of all stages for each species. In nonsevere malaria, mostly the young stages (<24

12
hours old) of P. falciparum are present in the peripheral blood, whereas for severe
malaria all stages can be present in the peripheral blood. For P. falciparum, the
trophozoite-infected red blood cells disappear from the peripheral blood circulation by
attachment to the walls of capillaries inside vital organs, which is a process called
sequestration. If the capillaries are blocked for newly infected cells by already
attached cells, more mature parasite stages (trophozoites and schizonts) will be visible
in the peripheral blood, which indicates a severe infection and a bad prognosis.
For P. falciparum, ring stages have a visible cytoplasm and 1 or 2 small chromatin
dots. The infected blood cells are not enlarged but can feature multiple infections. P.
falciparum trophozoites are rarely seen in peripheral blood smears. The cytoplasm of
mature trophozoites tends to be more dense than younger rings, trophozoites can
appear round in shape with brown malarial pigment inside, (Centers for Disease
Control and Prevention (CDC)). P. falciparum schizonts are also seldomly seen in
peripheral blood. They are displaying more than 2 and up to 32 nuclei (merozoites)
with dark brown pigment clumped in the middle. Gametocytes of P. falciparum have
a crescent or sausage shape, and can be seen in the blood smear 1 week after a
parasite infection. The chromatin is visible as a single mass or is diffuse. For more
information about P. falciparum morphology, see for example Similar observations
can be made for the stages of the other parasite species. For example, for P. vivax,
host cells are often enlarged and have irregular shape. Trophozoites are amoe
boid in shape with malaria pigment seen, and for severe infections multiple infections
of single blood cells are not uncommon. For P. malariae, host cells are not en
larged. Trophozoites have a strong tendency to form a band with malarial pigment
scattered along across the diameter of infected red blood cells. Multiple infec
tions are extremely rare for P. malariae. On the other hand, for P. ovale, host cells are
slightly enlarged and have an oval shape with tufted ends, often fimbriated.
Parasites are slightly enlarged and trophozoites are amoeboid in shape with malarial
pigment. Multiple infections of a single cell are more common than for P. vivax. For
P. knowlesi, infected red blood cells do not appear enlarged. The parasite erythocytic
cycle is only 24 hours, which is shorter than P. falciparum‘s cycle (48 hours) and
much shorter than P. malariae‘s cycle (72 hours), which will lead to the same stage
seen in peripheral blood every day at a given time. The morphology of P. knowlesi
parasites is similar to P. malariae. Trophozoites can feature malarial pigment spread
inside, band form may be seen like P. malariae, but their cytoplasm is more irregular,
and multiple parasites infecting 1 single red blood cell can be seen like in P.
falciparum. examples of different parasite stages in the same thin blood slide image.
In the first slide image, P. falciparum trophozoites and gametocytes can be seen
together with white blood cells. The latter are larger and have a pronounced nucleus
compared with the many red blood cells in the image. In the second image, P.
falciparum ring stages are together with schizonts. In addition, other objects such as
parasite outside cells and staining noise are visible in both images. Staining noise
in particular can be confused with parasites by an unexperienced microscopist.

MALARIA DIAGNOSIS

Malaria is a curable disease, with drugs available for treatment, including drugs that
can help prevent malaria infections in travelers to malaria-prone regions. However,
there exists no effective vaccine against malaria yet, although this is an area of active
research and field studies. Once infected, malaria is a rapidly progressing disease,

13
with a serious risk of developing into severe and cerebral malaria with neurologic
symptoms for P. falciparum infections. Therefore, a timely diagnosis of malaria is
very important. Although malaria can be diagnosed in many different ways, there is
room for improvement for current malaria diagnostic tests including reducing cost,
increasing specificity, and improving ease of use. Because automated malaria
diagnosis for resource-poor settings is the main topic of this survey, we have devoted
2 subsections to light microscopy and RDTs, which are by far the 2 most heavily used
diagnostic means in these areas. We also briefly discuss the other options for malaria
diagnosis, although they are arguably less suited for the conditions in remote malaria
regions. For more information about malaria diagnosis, we refer readers to the surveys
and the following references.Detecting the presence of parasites is the key to malaria
diagnosis. In addition, identifying the parasite species and presence of potentially
mixed infections is important, as well as the observation of the stage development of
P. falciparum parasites in relation to the severity of the disease. Counting parasites for
determining the level of parasitemia is not only important for identifying an in
fection and measuring its severity, it also allows monitoring patients by measuring
drug efficacy and potential drug resistance. Light microscopy. The current gold-
standard method for malaria diagnosis in the field is light microscopy of blood films,
which is the main focus of this article. Although other forms of diagnosis exist and
have become popular in recent years, in particular RDTs, microsco
py remains the most popular diagnostic tool, especially in resource-poor settings.
With microscopy, all parasite species can be detected. It allows computing the level
of parasitemia, clearing a patient after a successful treatment, and monitoring drug
resistance. Furthermore, it is less expensive than other methods and widely avail
able. However, its biggest disadvantages are the extensive training required for a
microscopist to become a proficient malaria slide reader, the high cost of training and
employing, maintaining skills, and the large component of manual work involved.
April 2018To diagnose malaria under a microscope, a drop of the patient‘s blood is
applied to a glass slide, which is then immersed in a staining solution to make
parasites more easily visible under a conventional light microscope, usually with a
100× oil objective. Two different types of blood smears are typically prepared for
malaria diagnosis: thick and thin smears.A thick smear is used to detect the presence
of parasites in a drop of blood. Thick smears allow a more efficient detection of
parasites than thin smears, with an 11 times higher sensitivity. On the other hand, thin
smears, which are the result of spreading the drop of blood across the glass slide, have
other advantages. They allow the examiner to identify malaria species and recognize
parasite stages more easily. The actual microscopic examination of a single blood
slide, including quantitative parasite detection and species identification, takes a
trained microscopist 15–30 minutes. Considering that hundreds of thousands of blood
slides are manually inspected for malaria every year, this amounts to a huge economic
effort required for malaria diagnosis. Rapid diagnostic tests. The main advantage of
microscopic malaria diagnosis lies in its low direct cost, which gives it a distinct
advantage in resource-poor settings.Other existing diagnostic methods, and any new
method, have to prove that they can provide the same ease of use and price point as
microscopy given the limited financial resources typically available in malaria-prone
regions. Arguably the only and main competitor in this sense are
RDTs. They detect evidence of malaria parasites (antigens) and take about 10–15
minutes to process. Their detection sensitivity is lower but comparable with manual
microscopy, and they do not require any special equipment and require only minimal
training. Although RDTs are currently more expensive than microscopy in high-

14
burden areas, a valid question is whether these tests can replace microscopy in the
near future. At the time of this writing, according to WHO,more countries use
microscopy more than they use RDTs.RDTs are used more in rural areas where
microscopy is not available. About 47% of malaria tests in malaria endemic countries
worldwide were made by RDT.The use of RDTs, however, does not eliminate the
need for malaria microscopy. A major disadvantage is that RDTs do not provide
quantification of the results. Therefore, at this point in time, microscopy and RDTs
are more complementing each other than one replacing the other. Other tests. Several
methods for diagnosing malaria are available. Important criteria are cost per test,
sensitivity and specificity of the method, time per test, and the required skill level of
the user. Furthermore, quantification of the number of infected red blood cells is
important as a prognostic indicator.
•Polymerase chain reaction (PCR). A molecular method called PCR has shown higher
sensitivity and specificity than conventional microscopic examination of stained
peripheral blood smears In fact, it is considered the most accurate among all tests. It
can detect very low parasite concentrations in the blood and can differentiate species.
However, PCR is a complex highcost technology that takes many hours to process by
trained staff. According to Tangpukdee et al., PCR is not routinely implemented in
developing countries because of the complexity of the testing and the lack of
resources to perform these tests adequately and routinely. Quality control and
equipment maintenance are also essential for the PCR technique, so that it may not be
suitable for malaria diagnosis in remote rural areas or even in routine clinical
diagnostic settings.
•Fluorescent microscopy. Quantitative buffy coat is a laboratory test to detect
infection with malaria or other blood parasites, using fluorescent microscopy. A
fluorescent dye makes parasites visible under ultraviolet light. According to Adeoye
and Nga, this test is more sensitive than the conventional thick smear. Nowadays,
portable fluorescent microscopes with fluorescent reagent to label parasites, are
available commercially. Although the quantitative buffy coat technique is simple,
reliable, and user friendly, it requires specialized instrumentation, is more costly than
conventional light microscopy, and is poor at determining species and numbers of
parasites.
•Flow cytometry. This is a laser-based cell counting and detection method that allows
to profile thousands of cells per second. Although flow cytometry offers automated
parasitemia counts, this is offset by a rather low sensitivity. Flow cytometry is less
suitable as a diagnostic technique in the field, when a direct answer is required for
treatment decisions. However, in developed countries, it can be applied in the clinical
setting for accurate counting of parasite numbers, for instance in the follow-up of drug
treatment.

STAINING METHODS

More than 100 years ago, Giemsas stain (1902) was applied for the first time for the
diagnosis of malaria. Since then, it received increased attention. Because of its low
cost, its high sensitivity, and specificity, it is currently widely used in microscopical
malaria examinations.However, Giemsa staining requires multiple reagents, ex
perienced personal, and is labor-intensive and timeconsuming (it typically requires at
least 45 minutes to stain a slide). Other stains have been used, too, like Field stain that
significantly reduces the staining time, although it requires drying of samples before
and during staining.However there are also disadvantages with Field‘s stain, specially

15