Malaria antigen detection tests are a group of commercially available

rapid diagnostic test A rapid diagnostic test (RDT) is a medical diagnostic test that is quick and easy to perform. RDTs are suitable for preliminary or emergency Screening (medicine), medical screening and for use in medical facilities with limited resources. They als ...

s of the

rapid antigen test A rapid antigen test (RAT), sometimes called a rapid antigen detection test (RADT), antigen rapid test (ART), or wikt:Appendix:Glossary#loosely, loosely just a rapid test, is a rapid diagnostic test suitable for point-of-care testing that directl ...

type that allow quick

diagnosis Diagnosis (: diagnoses) is the identification of the nature and cause of a certain phenomenon. Diagnosis is used in a lot of different academic discipline, disciplines, with variations in the use of logic, analytics, and experience, to determine " ...

malaria Malaria is a Mosquito-borne disease, mosquito-borne infectious disease that affects vertebrates and ''Anopheles'' mosquitoes. Human malaria causes Signs and symptoms, symptoms that typically include fever, Fatigue (medical), fatigue, vomitin ...

by people who are not otherwise skilled in traditional laboratory techniques for diagnosing malaria or in situations where such equipment is not available. There are currently over 20 such tests commercially available (WHO product testing 2008). The first malaria

antigen In immunology, an antigen (Ag) is a molecule, moiety, foreign particulate matter, or an allergen, such as pollen, that can bind to a specific antibody or T-cell receptor. The presence of antigens in the body may trigger an immune response. ...

suitable as target for such a test was a soluble

glycolytic Glycolysis is the metabolic pathway that converts glucose () into pyruvic acid, pyruvate and, in most organisms, occurs in the liquid part of cells (the cytosol). The Thermodynamic free energy, free energy released in this process is used to form ...

enzyme An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different mol ...

Glutamate dehydrogenase Glutamate dehydrogenase (GLDH, GDH) is an enzyme observed in both prokaryotes and eukaryotic mitochondria. The aforementioned reaction also yields ammonia, which in eukaryotes is canonically processed as a substrate in the urea cycle. Typic ...

. None of the rapid tests are currently as sensitive as a thick

blood film A blood smear, peripheral blood smear or blood film is a thin layer of blood smeared on a glass microscope slide and then stained in such a way as to allow the various blood cells to be examined microscopically. Blood smears are examined in the i ...

, nor as cheap. A major drawback in the use of all current dipstick methods is that the result is essentially qualitative. In many endemic areas of

tropical Africa The Afrotropical realm is one of the Earth's eight biogeographic realms. It includes Sub-Saharan Africa, the southern Arabian Peninsula, the island of Madagascar, and the islands of the western Indian Ocean. It was formerly known as the Ethiopi ...

, however, the quantitative assessment of parasitaemia is important, as a large percentage of the population will test positive in any qualitative assay.

Antigen-based Malaria Rapid Diagnostic Tests

Malaria is a curable

disease A disease is a particular abnormal condition that adversely affects the structure or function (biology), function of all or part of an organism and is not immediately due to any external injury. Diseases are often known to be medical condi ...

if the patients have access to early diagnosis and prompt treatment. Antigen-based

(RDTs)
have an important role at the periphery of health services capability because many rural

clinic A clinic (or outpatient clinic or ambulatory care clinic) is a health facility that is primarily focused on the care of outpatients. Clinics can be privately operated or publicly managed and funded. They typically cover the primary care needs ...

s do not have the ability to diagnose malaria on-site due to a lack of microscopes and trained

technician A technician is a worker in a field of technology who is proficient in the relevant skill and technique, with a relatively practical understanding of the theoretical principles. Specialisation The term technician covers many different special ...

s to evaluate blood films. Furthermore, in regions where the disease is not

endemic Endemism is the state of a species being found only in a single defined geographic location, such as an island, state, nation, country or other defined zone; organisms that are indigenous to a place are not endemic to it if they are also foun ...

, laboratory technologists have very limited experience in detecting and identifying malaria parasites. An ever increasing numbers of travelers from temperate areas each year visit tropical countries and many of them return with a malaria infection. The RDT tests are still regarded as complements to conventional

microscopy Microscopy is the technical field of using microscopes to view subjects too small to be seen with the naked eye (objects that are not within the resolution range of the normal eye). There are three well-known branches of microscopy: optical mic ...

but with some improvements it may well replace the

microscope A microscope () is a laboratory equipment, laboratory instrument used to examine objects that are too small to be seen by the naked eye. Microscopy is the science of investigating small objects and structures using a microscope. Microscopic ...

. The tests are simple and the procedure can be performed on the spot in field conditions. These tests use finger-stick or venous

blood Blood is a body fluid in the circulatory system of humans and other vertebrates that delivers necessary substances such as nutrients and oxygen to the cells, and transports metabolic waste products away from those same cells. Blood is com ...

, the completed test takes a total of 15–20 minutes, and a

laboratory A laboratory (; ; colloquially lab) is a facility that provides controlled conditions in which scientific or technological research, experiments, and measurement may be performed. Laboratories are found in a variety of settings such as schools ...

is not needed. The threshold of detection by these rapid diagnostic tests is in the range of 100 parasites/μL of blood compared to 5 by thick film microscopy.

pGluDH

Plasmodium glutamate dehydrogenase precipitation

An accurate diagnosis is becoming more and more important, in view of the increasing resistance of ''

Plasmodium falciparum ''Plasmodium falciparum'' is a Unicellular organism, unicellular protozoan parasite of humans and is the deadliest species of ''Plasmodium'' that causes malaria in humans. The parasite is transmitted through the bite of a female ''Anopheles'' mos ...

'' and the high price of alternatives to

chloroquine Chloroquine is an antiparasitic medication that treats malaria. It works by increasing the levels of heme in the blood, a substance toxic to the malarial parasite. This kills the parasite and stops the infection from spreading. Certain types ...

. The enzyme pGluDH does not occur in the host

red blood cell Red blood cells (RBCs), referred to as erythrocytes (, with -''cyte'' translated as 'cell' in modern usage) in academia and medical publishing, also known as red cells, erythroid cells, and rarely haematids, are the most common type of blood cel ...

and was recommended as a marker enzyme for ''Plasmodium'' species by Picard-Maureau et al. in 1975. The malaria marker enzyme test is suitable for routine work and is now a standard test in most departments dealing with malaria. Presence of pGluDH is known to represent parasite viability and a rapid diagnostic test using pGluDH as antigen would have the ability to differentiate live from dead organisms. A complete RDT with pGluDH as antigen has been developed in

China China, officially the People's Republic of China (PRC), is a country in East Asia. With population of China, a population exceeding 1.4 billion, it is the list of countries by population (United Nations), second-most populous country after ...

and is now undergoing

clinical trial Clinical trials are prospective biomedical or behavioral research studies on human subject research, human participants designed to answer specific questions about biomedical or behavioral interventions, including new treatments (such as novel v ...

s. GluDHs are ubiquitous enzymes that occupy an important branch-point between carbon and nitrogen metabolism. Both

nicotinamide adenine dinucleotide Nicotinamide adenine dinucleotide (NAD) is a Cofactor (biochemistry), coenzyme central to metabolism. Found in all living cell (biology), cells, NAD is called a dinucleotide because it consists of two nucleotides joined through their phosphat ...

(NAD) C 1.4.1.2and

nicotinamide adenine dinucleotide phosphate Nicotinamide adenine dinucleotide phosphate, abbreviated NADP or, in older notation, TPN (triphosphopyridine nucleotide), is a Cofactor (biochemistry), cofactor used in anabolic reactions, such as the Calvin cycle and lipid and nucleic acid synt ...

(NADP) dependent GluDH C 1.4.1.4enzymes are present in ''Plasmodia''; the NAD-dependent GluDH is relatively unstable and not useful for diagnostic purposes. Glutamate dehydrogenase provides an oxidizable carbon source used for the production of energy as well as a reduced electron carrier, NADH.

Glutamate Glutamic acid (symbol Glu or E; known as glutamate in its anionic form) is an α-amino acid that is used by almost all living beings in the biosynthesis of proteins. It is a Essential amino acid, non-essential nutrient for humans, meaning that ...

is a principal amino donor to other

amino acids Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although over 500 amino acids exist in nature, by far the most important are the Proteinogenic amino acid, 22 α-amino acids incorporated into p ...

in subsequent transamination reactions. The multiple roles of glutamate in nitrogen balance make it a gateway between free ammonia and the amino groups of most amino acids. Its

crystal structure In crystallography, crystal structure is a description of ordered arrangement of atoms, ions, or molecules in a crystalline material. Ordered structures occur from intrinsic nature of constituent particles to form symmetric patterns that repeat ...

is published. The GluDH activity in '' P.vivax'', '' P.ovale'' and '' P. malariae'' has never been tested, but given the importance of GluDH as a branch point enzyme, every cell must have a high concentration of GluDH. It is well known that enzymes with a high molecular weight (like GluDH) have many

isozyme In biochemistry, isozymes (also known as isoenzymes or more generally as multiple forms of enzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. Isozymes usually have different kinetic parameters (e.g. di ...

s, which allows strain differentiations (given the right

monoclonal antibody A monoclonal antibody (mAb, more rarely called moAb) is an antibody produced from a cell lineage made by cloning a unique white blood cell. All subsequent antibodies derived this way trace back to a unique parent cell. Monoclonal antibodie ...

). The host produces antibodies against the parasitic enzyme indicating a low sequence identity.

Histidine rich protein II

The histidine-rich protein II (HRP II) is a

histidine Histidine (symbol His or H) is an essential amino acid that is used in the biosynthesis of proteins. It contains an Amine, α-amino group (which is in the protonated –NH3+ form under Physiological condition, biological conditions), a carboxylic ...

- and

alanine Alanine (symbol Ala or A), or α-alanine, is an α-amino acid that is used in the biosynthesis of proteins. It contains an amine group and a carboxylic acid group, both attached to the central carbon atom which also carries a methyl group sid ...

-rich, water-soluble protein, which is localized in several cell compartments including the parasite cytoplasm. The antigen is expressed only by ''P. falciparum'' trophozoites. HRP II from ''P. falciparum'' has been implicated in the

biocrystallization Biocrystallization is the formation of crystals from organic macromolecules by living organisms. This may be a stress response, a normal part of metabolism such as processes that dispose of waste compounds, or a pathology. Template mediated crysta ...

hemozoin Haemozoin is a disposal product formed from the digestion of blood by some blood-feeding parasites. These hematophagy, hematophagous organisms such as malaria parasites (''Plasmodium spp.''), ''Rhodnius'' and ''Schistosoma'' digest haemoglobin an ...

, an inert, crystalline form of ferriprotoporphyrin IX (Fe(3+)-PPIX) produced by the parasite. A substantial amount of the HRP II is secreted by the parasite into the host bloodstream and the antigen can be detected in

erythrocytes Red blood cells (RBCs), referred to as erythrocytes (, with -''cyte'' translated as 'cell' in modern usage) in academia and medical publishing, also known as red cells, erythroid cells, and rarely haematids, are the most common type of blood cel ...

, serum, plasma,

cerebrospinal fluid Cerebrospinal fluid (CSF) is a clear, colorless Extracellular fluid#Transcellular fluid, transcellular body fluid found within the meninges, meningeal tissue that surrounds the vertebrate brain and spinal cord, and in the ventricular system, ven ...

and even

urine Urine is a liquid by-product of metabolism in humans and many other animals. In placental mammals, urine flows from the Kidney (vertebrates), kidneys through the ureters to the urinary bladder and exits the urethra through the penile meatus (mal ...

as a secreted water-soluble protein. These antigens persist in the circulating blood after the parasitaemia has cleared or has been greatly reduced. It generally takes around two weeks after successful treatment for HRP2-based tests to turn negative, but may take as long as one month, which compromises their value in the detection of active infection. False positive dipstick results were reported in patients with rheumatoid-factor-positive

rheumatoid arthritis Rheumatoid arthritis (RA) is a long-term autoimmune disorder that primarily affects synovial joint, joints. It typically results in warm, swollen, and painful joints. Pain and stiffness often worsen following rest. Most commonly, the wrist and h ...

. Since HRP-2 is expressed only by ''P. falciparum'', these tests will give negative results with samples containing only ''P. vivax'', ''P. ovale'', or ''P. malariae''; many cases of non-falciparum malaria may therefore be misdiagnosed as malaria negative (some ''P.falciparum'' strains also don't have HRP II). The variability in the results of pHRP2-based RDTs is related to the variability in the target antigen.

pLDH

''P. falciparum''

lactate dehydrogenase Lactate dehydrogenase (LDH or LD) is an enzyme found in nearly all living cells. LDH catalyzes the conversion of pyruvic acid, pyruvate to lactic acid, lactate and back, as it converts NAD+ to NADH and back. A dehydrogenase is an enzyme that t ...

(PfLDH) is a 33 kDa

oxidoreductase In biochemistry, an oxidoreductase is an enzyme that catalyzes the transfer of electrons from one molecule, the reductant, also called the electron donor, to another, the oxidant, also called the electron acceptor. This group of enzymes usually ut ...

C 1.1.1.27 It is the last enzyme of the glycolytic pathway, essential for ATP generation and one of the most abundant enzymes expressed by ''P. falciparum''. ''Plasmodium'' LDH (pLDH) from ''P. vivax'', ''P. malariae'', and ''P. ovale'') exhibit 90-92% identity to PfLDH from ''P. falciparum''. pLDH levels have been seen to reduce in the blood sooner after treatment than HRP2. In this respect, pLDH is similar to pGluDH. Nevertheless, the kinetic properties and sensitivities to inhibitors targeted to the cofactor binding site differ significantly and are identifiable by measuring dissociation constants for inhibitors which, differ by up to 21-fold.

pAldo

Fructose-bisphosphate aldolase Fructose-bisphosphate aldolase (), often just aldolase, is an enzyme catalyzing a reversible reaction that splits the aldol, fructose 1,6-bisphosphate, into the triose phosphates dihydroxyacetone phosphate (DHAP) and glyceraldehyde 3-phosp ...

C 4.1.2.13catalyzes a key reaction in

glycolysis Glycolysis is the metabolic pathway that converts glucose () into pyruvic acid, pyruvate and, in most organisms, occurs in the liquid part of cells (the cytosol). The Thermodynamic free energy, free energy released in this process is used to form ...

and energy production and is produced by all four species. The ''P.falciparum'' aldolase is a 41 kDa protein and has 61-68% sequence similarity to known

eukaryotic The eukaryotes ( ) constitute the Domain (biology), domain of Eukaryota or Eukarya, organisms whose Cell (biology), cells have a membrane-bound cell nucleus, nucleus. All animals, plants, Fungus, fungi, seaweeds, and many unicellular organisms ...

aldolases. Its crystal structure has been published. The presence of antibodies against p41 in the sera of human adults partially immune to malaria suggest that p41 is implicated in protective immune response against the parasite.

References

External links

Malaria Antibodies

WHO product testing 2008

WHO Rapid Diagnostic Tests (RDTs)
{{malaria Malaria Blood tests