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International Council for Standardization in Haematology (ICSH)

Published in Hemotology
Saturday, 22 July 2017 06:23
The International Council for Standardization in Haematology (ICSH) was initiated as a standardization committee by the European Society of Haematology (ESH) in 1963 and officially constituted by the International Society of Hematology (ISH) and the ESH in Stockholm in 1964. The ICSH is recognised as a Non-Governmental Organisation with official relations to the World Health Organisation (WHO).
The ICSH is a not-for-profit organisation that aims to achieve reliable and reproducible results in laboratory analysis in the field of diagnostic haematology.
The ICSH coordinates Working Groups of experts to examine laboratory methods and instruments for haematological analyses, to deliberate on issues of standardization and to stimulate and coordinate scientific work as necessary towards the development of international standardization materials and guidelines.

Prickly Heat Rash: Cause, Symptoms and Treatment

Published in Informative
Sunday, 02 July 2017 10:39
Prickly heat usually clears up on its own within a few days. However, in serious cases heat rash can interfere with the body's heat-regulating mechanism and cause heat exhaustion.
Heatstroke is a more serious condition when the body can no longer cool itself. This is a medical emergency.
What causes prickly heat rash?
Heat rash begins with excessive perspiration, usually in a hot, humid environment. The perspiration makes it easier for dead skin cells and bacteria on the skin to block the sweat glands, forming a barrier and trapping sweat beneath the skin, where it builds up, causing the characteristic bumps. As the bumps burst and sweat is released, there may be a prickly, or stinging sensation that gives this condition its name.

What are the symptoms of heat rash?
Small, itchy red bumps on the skin are the symptoms of heat rash. The rash may feel prickly, stinging or burning.
Seek medical advice if:
  • Heat rash does not go away on its own within a few days.
  • You develop an infection in an area where you recently had heat rash.
What are the treatments for heat rash?
In most cases, heat rash will clear up on its own in a few days if the affected area is kept cool and dry. Avoid excessive heat and humidity and cool off with a fan, take a cool shower or bath and let your skin air dry, or if you have air-conditioning, use this to cool yourself. Once the skin is cool and dry again, don’t use any type of oil-based product, which might block your sweat glands. Calamine lotion and/or hydrocortisone cream can relieve itching and irritation.
If your prickly heat does not go away within a few days, or if you develop an infection where the bumps have burst, you may need medication, so seek medical advice.

How can I prevent heat rash?
To prevent heat rash, avoid situations that can lead to excessive sweating, such as hot, humid environments and strenuous physical activity. In hot weather, use fans and cool showers and baths to stay cool, or air conditioning if available; dry your skin thoroughly; and wear lightweight, loose-fitting clothes ideally made from cotton.


Published in Informative
Monday, 26 June 2017 19:27
  • Allen’s LawAllen
  • Artificial ParthenogenesisLoeb
  • Axial Gradient theoryChild
  • Bergman’s RuleBergman
  • Biogenetic LawEarnst Haeckel (1868)
  • Biological Species ConceptEarnst Mayer
  • Biogenesis TheoryDeveloped by F. Redi
  • Chromosomal Theory of InheritanceSutton and Boveri
  • Theory of natural selectionCharles Darwin


Published in Informative
Monday, 26 June 2017 18:39

Amino acid sequence of protein (insulin)

Anaerobic release of energy
L-Pasteur (1878)


Pure culture of Bacteria
Lister J.

Towrt and De Herelle (1915)

Blood Capillaries
Marcello Malpighi

Blood Groups
Karl Landsteiner

Blood Circulation
William Harvey

E.R. Dubois


Saffermann and Morris

First description of cell (RBC)
Jan Swammerdam (1658)

Robert Hooke (1665)

Living cell
A.V. Leeuwenhoek

Cell Theory
Schleiden and Schwann

Van Benden

Van Benden


Golgi bodies
Cammileo Golgi

Haeckel (1866)


Kolliker (1880)

Robertis and Francis

Paleviz et. al (1975)

Robert Brown



Ribosomes (Animal cell)

Pernes (1953)

Astral rays and spindle

Endoplasmic reticulum

Central Dogma
F.H.C. Crick (1918)

Coenzyme A
C. Lipmann

Chlorophyll structure
Willstartter and Fisher


C.A. Macmunn (1886)

Citric Acid cycle
Hans A. Krebs

Double Helical Structure of DNA
Watson and Crick

Biological Synthesis of DNA with template
A. Kornberg.

biological synthesis of DNA without template
H.G. Khorona


Embryo culture

Extra embryonic membranes
Von Baer

Fertilization in plants
E. Strasburger

Double fertilization

Go phase

Gaseous exchange in blood
Ludwig (1872)

Genetic defects in human
Sir Archibald Garrod

Giant Salivary gland chromosomes
Balbiani (1881)

Beylis and Starling


Issacs and Linderman

Insulin use for treatment of diabetics

G. Mendel

Rediscoverer of Mendelism
Correns, Hugo de Vries and Tschmark

W. His


W. Flemming

Farmer and Moore

Hugo de Vries

Nucleic acid
Meishcher called it ‘Nuclein’

Ovum (Mammalian)
Karl E. Von Baer

Omnis cellula e cellula
R. Virchow

Edward and Lewis


Alexander Flemming


Garner and Allard

Park and Bigginis (1960)

Quiescent centre

Protoplasm Physical basis of life

Salmon Waksman

Techniques Chromatograph
M. Tswett

Tissue culture
A. Carrel

Isotopic tracing
G. Havesy

Measuring gaseous exchange manometry
O. Warburg

Locating DNA in cell
A. Feulgen

T. Svedberg

Avena curvature test

Teminism (Reverse Transcription)

Synthesis of urea

D. Iwanovsky

Obtained crystals of virus


Published in Informative
Sunday, 25 June 2017 17:05

Antileprosy day
30th Jan

International Women day
8th March

World Handicaps day
15th March

World Forest day
21st March

World Tuberculosis day
26th March

World Health day
7th April

World Earth day
22nd April

International Sun day (Non-conventional Energy sources day)
3rd May

World Red Cross day
8th May

World No Tobacco Day
31st May

World Environment day
5th June

World Population Day
11th July

Hiroshima and Nagasaki Day
6th Aug

Malaria day (mosquito day)
20th Aug

Blood Donation Day
1st Oct

World Animal Day
3rd Oct

World habitat Day
4th Oct

World Food Day
16th Oct

World Diabetes Day
14 Nov

World AIDS Day
1st Dec

National Pollution Prevention Day
2nd Dec

World conservation Day
3rd Dec

International Day for Biological Diversity
29th Dec


Published in Informative
Friday, 23 June 2017 19:25
  • Abdul Qadir Khan Research Laboratories
  • Energy Conservation Cell (ENERCON), Islamabad
  • Drainage Research Institute of Pakistan (DRIP), Hyderabad
  • Forestry Institute, Peshawar
  • Ghulam Ishaq Khan Institute of Advanced Science and Technology, Tarbella
  • Geological Survey of Pakistan, Rawalpindi
  • Irrigation, Drainage, and Flood Control Research Council, Islamabad
  • National Center for Technology Transfer (NCTT), Islamabad
  • National Institute of Health (NIH), Islamabad
  • Nuclear Institute of Agricultural Biology (NIAB), Faisalabad
  • Pakistan Agricultural Research Council (PARC), Islamabad
  • Pakistan Arts Council
  • Pakistan Atomic Energy Commission (PAEC), Islamabad
  • Pakistan Council of Industrial and Scientific Research (PCSIR)
  • Pakistan Science Foundation (PSF), Islamabad
  • Pakistan Health Research Council (PHRC), Islamabad
  • Silicon Institute of Technology, Islamabad
  • Space and Upper Atmosphere Research Council (SUPPARCO), Karachi

H. Gobind Khorana - Biographical

Published in Informative
Sunday, 12 March 2017 13:08

Har Gobind Khorana was born of Hindu parents in Raipur, a little village in Punjab, which is now part of eastern Pakistan. The correct date of his birth is not known; that shown in documents is January 9th, 1922. He is the youngest of a family of one daughter and four sons. His father was a «patwari», a village agricultural taxation clerk in the British Indian system of government. Although poor, his father was dedicated to educating his children and they were practically the only literate family in the village inhabited by about 100 people.

Har Gobind Khorana attended D.A.V. High School in Multan (now West Punjab); Ratan Lal, one of his teachers, influenced him greatly during that period. Later, he studied at the Punjab University in Lahore where he obtained an M. Sc. degree. Mahan Singh, a great teacher and accurate experimentalist, was his supervisor.

Khorana lived in India until 1945, when the award of a Government of India Fellowship made it possible for him to go to England and he studied for a Ph. D. degree at the University of Liverpool. Roger J. S. Beer supervised his research, and, in addition, looked after him diligently. It was the introduction of Khorana to Western civilization and culture.

Khorana spent a postdoctoral year (1948-1949) at the Eidgenössische Technische Hochschule in Zurich with Professor Vladimir Prelog. The association with Professor Prelog molded immeasurably his thought and philosophy towards science, work, and effort.

After a brief period in India in the fall of 1949, Khorana returned to England where he obtained a fellowship to work with Dr. (now Professor) G. W. Kenner and Professor (now Lord) A. R. Todd. He stayed in Cambridge from 1950 till 1952. Again, this stay proved to be of decisive value to Khorana. Interest in both proteins and nucleic acids took root at that time.

A job offer in 1952 from Dr. Gordon M. Shrum of British Columbia (now Chancellor of Simon Fraser University, British Columbia) took him to Vancouver. The British Columbia Research Council offered at that time very little by way of facilities, but there was «all the freedom in the world», to use Dr. Shrum's words, to do what the researcher liked to do. During the following years, with Dr. Shrum's inspiration and encouragement and frequent help and scientific counsel from Dr. Jack Campbell (now Head of the Department of Microbiology at the University of British Columbia), a group began to work in the field of biologically interesting phosphate esters and nucleic acids. Among the many devoted and loyal colleagues of this period, there should, in particular, be mention of Dr. Gordon M. Tener (now a Professor in the Biochemistry Department of the University of British Columbia), who contributed much to the spiritual and intellectual well-being of the group.

In 1960 Khorana moved to the Institute for Enzyme Research at the University of Wisconsin. He became a naturalized citizen of the United States. As of the fall of 1970 Khorana has been Alfred P. Sloan Professor of Biology and Chemistry at the Massachusetts Institute of Technology.

Har Gobind Khorana was married in 1952 to Esther Elizabeth Sibler, who is of Swiss origin. Esther brought a consistent sense of purpose into his life at a time when, after six years' absence from the country of his birth, Khorana felt out of place everywhere and at home nowhere. They have three children: Julia Elizabeth (born May 4th, 1953), Emily Anne (born October 18th, 1954), and Dave Roy (born July 26th, 1958).

Towards a cure for herpesviruses: Targeting infection with CRISPR/Cas9

Published in Informative
Tuesday, 22 November 2016 08:35

Most adults carry multiple herpesviruses. Following the initial acute infection, these viruses establish life-long infections in their hosts and cause cold sores, keratitis, genital herpes, shingles, infectious mononucleosis, and other diseases. Some herpesviruses can cause cancer in man. During the latent phase of infection, the viruses remain dormant for long periods of time, but retain the capacity to cause occasional reactivations, that may lead to disease. A study published on June 30th in PLOS Pathogens suggests that attacking herpesvirus DNA with CRISPR/Cas9 genome editing technology can suppress virus replication and, in some cases, lead to elimination of the virus.

The CRISPR/Cas9 system targets specific DNA sequences and induces clean cuts across both strands of the DNA. In mammalian cells, such cuts are flagged and quickly repaired by an emergency repair system called NHEJ (for non-homologous end-joining). NHEJ is efficient but not very accurate and often results in insertion or deletion of a few DNA bases at the repair site. Because DNA is read in codons of three bases at a time, such small changes in critical positions often destroy the function of the respective gene and its protein product.

Robert Jan Lebbink, from the University Medical Center in Utrecht, The Netherlands, and colleagues reasoned that CRISPR/Cas9 could target and mutate latent herpesvirus DNA in infected human cells and so potentially prevent herpesvirus-associated diseases. To test this, the researchers devised specific guide (g)RNAs—sequences that are complementary to vital parts of the viral genome and function as 'molecular addresses'. These gRNAs, combined with the 'molecular scissors' part of the CRISPR/Cas9 system, should induce specific cuts and subsequent mutations in the herpesvirus DNA, and so cripple the viruses.

In their systematic approach, the researchers looked at three different members of the herpesvirus group: herpes simplex virus type 1 (HSV-1) causing cold sores and herpes keratitis; human cytomegalovirus (HCMV), the most common viral cause of birth defects (when the virus is transmitted from mother to fetus); and Epstein-Barr virus (EBV) causing infectious mononucleosis and multiple types of cancer.

Working with lymphoma cells latently infected with EBV, the researchers showed that introduction of gRNAs that target specific EBV DNA sequences can introduce mutations at the targeted sites. Such mutations can eliminate essential functions of the virus as well as de-stabilize the viral DNA molecules. Consistent with this, the researchers report that by using two different gRNAs targeting an essential EBV gene, they can induce loss of over 95% of EBV genomes from the host cells.

During latent infection, HCMV genomes exist as circular DNA molecules in the nucleus of host cells. Upon virus reactivation, HCMV replication proceeds slowly. With appropriate gRNAs, the researchers found that CRISPR/Cas9 editing can efficiently impair HCMV replication. However, they also observed emergence of escape variants that bypass CRISPR/Cas9 editing, suggesting that simultaneous editing at multiple critical sites in the HCMV genome is necessary to avoid the development of resistant genomes.

Compared to HCMV, HSV-1 multiplies much faster. When the researchers tested various gRNAs targeting different essential HSV-1 genes in conjunction with CRISPR/Cas9, they found that many of them were able to reduce virus replication. When they combined two of those gRNAs, thereby simultaneously targeting two essential genes, they were able to completely suppress HSV-1 replication. On the other hand, they were unable to induce editing during the latent phase, i.e. when the viral DNA was not actively multiplying.

"We observed highly efficient and specific clearance of EBV from latently infected tumor cells and impairment of HSV-1 and HCMV replication in human cells", the researchers summarize. They go on to say, "although CRISPR/Cas9 was inefficient at directing genome engineering of quiescent HSV-1, virus replication upon reactivation of quiescent HSV-1 was efficiently abrogated using anti-HSV-1 gRNAs". Their results, they hope, "may allow the design of effective therapeutic strategies to target human herpesviruses during both latent and productive infections."

Top 9 Deadliest Viruses on Earth

Published in Microbiology
Saturday, 29 October 2016 19:31

Deadly Diseases

Humans have been battling viruses since before our species had even evolved into its modern form. For some viral diseases, vaccines and antiviral drugs have allowed us to keep infections from spreading widely, and have helped sick people recover. For one disease — smallpox — we've been able to eradicate it, ridding the world of new cases.
But as the Ebola outbreak now devastating West Africa demonstrates, we're a long way from winning the fight against viruses.
The strain that is driving the current epidemic, Ebola Zaire, kills up to 90 percent of the people it infects, making it the most lethal member of the Ebola family. "It couldn't be worse," said Elke Muhlberger, an Ebola virus expert and associate professor of microbiology at Boston University.
But there are other viruses out there that are equally deadly, and some that are even deadlier. Here are the nine worst killers, based on the likelihood that a person will die if they are infected with one of them, the sheer numbers of people they have killed, and whether they represent a growing threat.

1. Marburg virus

Scientists identified Marburg virus in 1967, when small outbreaks occurred among lab workers in Germany who were exposed to infected monkeys imported from Uganda. Marburg virus is similar to Ebola in that both can cause hemorrhagic fever, meaning that infected people develop high fevers and bleeding throughout the body that can lead to shock, organ failure and death.
The mortality rate in the first outbreak was 25 percent, but it was more than 80 percent in the 1998-2000 outbreak in the Democratic Republic of Congo, as well as in the 2005 outbreak in Angola, according to the World Health Organization (WHO).

2. Ebola virus

The first known Ebola outbreaks in humans struck simultaneously in the Sudan and the Democratic Republic of Congo in 1976. Ebola is spread through contact with blood or other body fluids, or tissue from infected people or animals. The known strains vary dramatically in their deadliness, Muhlberger said.
One strain, Ebola Reston, doesn't even make people sick. But for the Bundibugyo strain, the fatality rate is up to 50 percent, and it is up to 71 percent percent for the Sudan strain, according to WHO.
The outbreak underway in West Africa began in early 2014, and is the largest and most complex outbreak of the disease to date, according to WHO.

3. Rabies

Although rabies vaccines for pets, which were introduced in the 1920s, have helped make the disease exceedingly rare in the developed world, this condition remains a serious problem in India and parts of Africa.
"It destroys the brain, it's a really, really bad disease," Muhlberger said. "We have a vaccine against rabies, and we have antibodies that work against rabies, so if someone gets bitten by a rabid animal we can treat this person," she said.
However, she said, "if you don't get treatment, there's a 100 percent possibility you will die."

4. HIV

In the modern world, the deadliest virus of all may be HIV. "It is still the one that is the biggest killer," said Dr. Amesh Adalja, an infectious disease physician and spokesman for the Infectious Disease Society of America.
An estimated 36 million people have died from HIV since the disease was first recognized in the early 1980s. "The infectious disease that takes the biggest toll on mankind right now is HIV," Adalja said.
Powerful antiviral drugs have made it possible for people to live for years with HIV. But the disease continues to devastate many low- and middle-income countries, where 95 percent of new HIV infections occur. Nearly 1 in every 20 adults in Sub-Saharan Africa is HIV-positive, according to WHO.

5. Smallpox

In 1980, the World Health Assembly declared the world free of smallpox. But before that, humans battled smallpox for thousands of years, and the disease killed about 1 in 3 of those it infected. It left survivors with deep, permanent scars and, often, blindness.
Mortality rates were far higher in populations outside of Europe, where people had little contact with the virus before visitors brought it to their regions.  For example, historians estimate 90 percent of the native population of the Americas died from smallpox introduced by European explorers. In the 20th century alone, smallpox killed 300 million people.
"It was something that had a huge burden on the planet, not just death but also blindness, and that's what spurred the campaign to eradicate from the Earth," Adalja said.

6. Hantavirus

Hantavirus pulmonary syndrome (HPS) first gained wide attention in the U.S. in 1993, when a healthy, young Navajo man and his fiancée living in the Four Corners area of the United States died within days of developing shortness of breath. A few months later, health authorities isolated hantavirus from a deer mouse living in the home of one of the infected people. More than 600 people in the U.S. have now contracted HPS, and 36 percent have died from the disease, according to the Centers for Disease Control and Prevention.
The virus is not transmitted from one person to another, rather, people contract the disease from exposure to the droppings of infected mice.
Previously, a different hantavirus caused an outbreak in the early 1950s, during the Korean War, according to a 2010 paper in the journal Clinical Microbiology Reviews. More than 3,000 troops became infected, and about 12 percent of them died.
While the virus was new to Western medicine when it was discovered in the U.S., researchers realized later that Navajo medical traditions describe a similar illness, and linked the disease to mice.

7. Influenza

During a typical flu season, up to 500,000 people worldwide will die from the illness, according to WHO. But occasionally, when a new flu strain emerges, a pandemic results with a faster spread of disease and, often, higher mortality rates.
The most deadly flu pandemic, sometimes called the Spanish flu, began in 1918 and sickened up to 40 percent of the world's population, killing an estimated 50 million people.
"I think that it is possible that something like the 1918 flu outbreak could occur again," Muhlberger said. "If a new influenza strain found its way in the human population,and could be transmitted easily between humans, and caused severe illness, we would have a big problem."

8. Dengue

Dengue virus first appeared in the 1950s in the Philippines and Thailand, and has since spread throughout the tropical and subtropical regions of the globe. Up to 40 percent of the world's population now lives in areas where dengue is endemic, and the disease — with the mosquitoes that carry it — is likely to spread farther as the world warms.
Dengue sickens 50 to 100 million people a year, according to WHO. Although the mortality rate for dengue fever is lower than some other viruses, at 2.5 percent, the virus can cause an Ebola-like disease called dengue hemorrhagic fever, and that condition has a mortality rate of 20 percent if left untreated.
"We really need to think more about dengue virus because it is a real threat to us," Muhlberger said. There is no current vaccine against dengue, but large clinical trials of an experimental vaccine developed by French drug maker Sanofi have had promising results.

9. Rotavirus

Two vaccines are now available to protect children from rotavirus, the leading cause of severe diarrheal illness among babies and young children. The virus can spread rapidly, through what researchers call the fecal-oral route (meaning that small particles of feces end up being consumed).
Although children in the developed world rarely die from rotavirus infection, the disease is a killer in the developing world, where rehydration treatments are not widely available.
The WHO estimates that worldwide, 453,000 children younger than age 5 died from rotavirus infection in 2008. But countries that have introduced the vaccine have reported sharp declines in rotavirus hospitalizations and deaths.
Author: Anne Harding, Contributing Writer
Source: Live Science

Preanalytical Errors in Clinical Chemistry Laboratory

Published in Biochemistry
Friday, 21 October 2016 17:57
Abstract: There are many factors that contribute to accurate test results in the chemistry laboratory. These factors can be broken down into three areas: preanalytical, analytical and post analytical. Preanalytical variables account for 32-75% of laboratory errors, and encompass the time from when the test is ordered by the physician until the sample is ready for analysis.1 The focus of this article will be preanalytical variables that can occur during a venipuncture and specimen processing and how they relate to testing in the clinical chemistry laboratory.
Scenario: A patient has been in the cardiac intensive care unit for 3 days. For the past 2 mornings, he has had his cardiac enzymes drawn into a BD SSTT tube to monitor his condition since his heart attack. On this particular morning, his tube of blood is drawn and sent to the clinical chemistry lab for analysis. However, when the tube is processed and ready for analysis, the technologist running the chemistry analyzer notices that the specimen is very gelatinous and will need to be re-processed before the sample can be run on the analyzer. What could have happened to the quality of this specimen?
There are many variables that can contribute to the quality of a chemistry specimen. This article will investigate the variables that may have contributed to the gelatinous specimen in the case of the cardiac patient, as well as the other variables that are important to specimen quality. The focus will be on the preanalytical phase of the blood collection and sample handling, up until the time that the sample is to be run on the chemistry instrument.
Following the above BD SST™ tube from time of collection until it is ready for analysis, the preanalytical variables that can contribute to the quality of the sample are as follows:
Patient Identification: It is important to identify a patient properly so that blood is being collected from the correct person. Drawing blood from the wrong person, or labeling the correct patient’s sample with a different patient’s label can certainly contribute to laboratory error. Perhaps in the opening scenario, the patient in the next bed, with an extremely prolonged clotting time, was drawn and labeled as the cardiac patient.
When identifying the patient, have them provide their full name, address, identification number and/or date of birth.2 Hospital inpatients should be wearing an identification band with the above information, which the phlebotomist should confirm before the venipuncture. Blood should not be drawn from a patient without a band. A nurse, physician, relative or guardian should identify patients that are unable to speak or identify themselves.
Patient Preparation: Prior to collecting specimens for chemistry, certain patient variables need to be considered. For certain chemistry analytes, such as glucose and cholesterol, patients need to be fasting (absence of food and liquids) for at least 12 hours prior to venipuncture. Other analytes, such as cortisol and adrenocorticotropin, have diurnal variations, where the analyte is at its highest level in the morning, and the levels gradually decrease during the course of the day.
Selecting the Site: Selecting the appropriate site for venipuncture can contribute to a better quality sample. The preferred site is the median cubital vein. This vein is usually the easiest to access. Generally, there is less need to probe to find the vein, which in turn should cause less trauma during the venipuncture. This will usually be the most comfortable for the patient. If the median cubital vein cannot be used, the next choice would be the cephalic vein. The last vein to consider for venipuncture is the basilic vein. This vein is in close proximity to the median nerve and brachial artery, and extreme caution must be used so that only the basilic vein is being punctured.
Site Preparation: Prior to venipuncture, the site should be cleansed with alcohol. Cleansing starts at the center of the vein, and should continue outward in concentric circles. Before performing the venipuncture, the alcohol should be allowed to air dry. This will help to ensure that the specimen is not contaminated with alcohol, as this can lead to hemolysis. Hemolysis can result in the spurious elevation of such analytes as potassium, lactate dehydrogenase (LD), iron and magnesium in the chemistry lab. Allowing the alcohol to dry completely will also cause less burning and pain to the patient.
Tourniquet Application and Time: The tourniquet should be applied approximately three to four inches above the venipuncture site. The tourniquet should be on the arm no longer than one minute. 2 A good rule of thumb to determine the one-minute tourniquet time is to remove the tourniquet when blood starts to flow into the first tube of blood being drawn. Prolonged tourniquet time can lead to an increase in various chemistry analytes, including serum protein, potassium and lactic acid due to hemoconcentration of blood at the puncture site.
Proper Venipuncture Technique: During phlebotomy, avoid probing to find the vein and achieve blood flow. Excessive probing and/or “fishing” to find a vein can result in a poor quality sample, including hemolysis. As mentioned previously, hemolysis can affect several chemistry analaytes.
Order of Draw: Following the correct order of draw during venipuncture will help to ensure accurate test results. The BD and CLSI (Clinical and Laboratory Standards Institute, formerly NCCLS) recommended order of draw for evacuated blood collection tubes is as follows (PDF).2
An example of improper order of draw that can lead to an incorrect chemistry result is drawing an EDTA tube prior to an BD SST ™ or heparin tube for chemistry testing. The potential cross contamination of K2 or K3EDTA on the needle from the lavender top tube to the chemistry tube can lead to an elevated potassium result. This in turn can require a recollection of the sample, or possible misdiagnosis or treatment of the patient.
Proper Tube Mixing: All tubes with additives need to be inverted to mix the additive evenly with the blood. Plastic serum tubes and BD SST™ tubes contain clot activator and should be inverted 5 times to mix the activator with the blood and help the specimen clot completely. In the opening case study, improper mixing of the tube after venipuncture could have contributed to the gelatinous serum sample that was seen in the laboratory. Other additive tubes, such as heparin, need to be inverted 8-10 times to mix the anticoagulant with the blood and prevent clotting. Be sure that tubes are not being shaken vigorously, as this can lead to a hemolyzed sample.
Correct Specimen Volume: All blood collection tubes need to be filled to the correct volume.3 This will ensure the proper amount of blood for the amount of additive in the tube (blood to additive ratio). For example, if a 5 mL draw heparin tube is only filled with 3 mL of blood, the heparin concentration is erroneously high and may potentially interfere with some chemistry analytes. Expiration dates should also be checked on the evacuated tubes.4 Expired tubes should not be used, as they may have a decreased vacuum, as well as potential changes in any additives in the tubes.
Proper Tube Handling and Specimen Processing: Once the blood collection tubes have been drawn in the correct order, to the proper fill volume and mixed thoroughly, the next step toward accurate test results is processing the tubes properly. This section will look at serum and plasma tubes separately, as both specimen types have their own special handling requirements.
Serum Samples
Serum specimens, namely red top tubes and BD SST™ gel tubes, need to clot completely prior to centrifugation and processing. Blood specimens in red top tubes should clot for 45 to 60 minutes, and those in BD SST ™ tubes should be allowed to clot for 30 minutes to ensure complete clot formation.4 Blood from patients who are receiving anticoagulant therapy, such as heparin or coumadin, may take longer to clot. Tubes should be allowed to clot at room temperature, upright in a test tube rack, with the closures on the tubes. In the gelatinous sample that was presented at the beginning of this article, perhaps the blood was not clotted completely prior to centrifugation because a cardiac patient is often heparanized. Spinning the tube too soon may result in a gelatinous and/or fibrinous serum sample that will require respinning.
Plasma Samples
Blood specimens collected in plasma tubes, such as the plain heparinized green top tubes and the BD PST™ tubes with heparin and gel do not require clotting prior to centrifugation. This allows the tube of blood to be drawn, mixed and centrifuged immediately, resulting in a quicker turn-around-time for test results.
Centrifugation: The next step in sample processing is the centrifugation of the blood collection tubes. Both BD SST™ and BD PST™ tubes are centrifuged at the same speed and for the same amount of time. In a swinging bucket centrifuge (preferred type of spin for gel separation tubes), the tubes should be spun for ten minutes at a speed of 1100 to 1300 relative centrifugal force (RCF). A fifteen-minute spin at the same speed is required for spinning tubes in a fixed- angle centrifuge. Serum and plasma tubes without gel can be spun at a speed of 1000 RCF for ten minutes.4
It is important to spin gel tubes for the recommended time. The gel barrier in the tubes needs time to move and form a solid barrier between the red cells and the serum or plasma. Also, in BD PST™ tubes, the white blood cells and platelets that remain in the plasma need adequate time to spin out of the plasma. If the BD PST™ tubes are spun for less than the recommended 10 minutes, these cells and platelets may remain in the plasma and could cause interference with some chemistry analytes. It is recommended that BD SST™ tubes should not be re-centrifuged after their initial centrifugation. Re-spinning the tubes can result in elevated potassium values, as excess serum that has been in contact with the red cells will be expressed from underneath the gel barrier.
Special Handling of Blood Specimens: Certain chemistry analytes will require the tube of blood to be chilled after collection in order to maintain the stability of the analyte. A slurry of ice and water is recommended for chilling the tubes of blood. Examples of specimens that need to be chilled or transported on ice include adrenocorticotropic hormone (ACTH), angiotensin converting enzyme (ACE), acetone, ammonia, catecholamines, free fatty acids, lactic acid, pyruvate and renin.
Other anayltes are photo-sensitive, and need to be protected from light in order to remain stable and to ensure that the laboratory reports an accurate result. This can be done by wrapping the tube of blood in aluminum foil. The most common example of a light-sensitive analyte is bilirubin. Other chemistry analytes that need to be light-protected include beta-carotene and erythrocyte protoporphyrin.
Stability for Whole Blood, Serum and Plasma: A whole blood specimen that is going to be spun down should be centrifuged and the serum or plasma removed from the red blood cells within two hours after the venipuncture.5 Once the serum has been removed or separated from the red blood cells (in the case of a gel barrier tube), the sample will be stable at room temperature for eight hours, and up to 48 hours at 2-4 degrees C.5 After 48 hours, the serum specimen should be frozen at –20 degrees C in an aliquot tube.5
Paying close attention to the preanalytical variables associated with blood collection will help to ensure accurate test results in the chemistry department, as well as all areas of the clinical laboratory. As was evident from the opening case study, there are often several variables that can potentially contribute to erroneous test results. Our cardiac patient’s blood could have been drawn from the wrong patient, had improper tube handling or his blood may have not clotted long enough. Therefore, it is important to remember that a better quality sample during the preanalytical phase of blood collection will yield a better test result.
1. Bonini P, PlebaniM, Ceriotti F, et al. Errors in laboratory medicine. Clin Chem. 2002;48:691-698.
2. NCCLS – Procedures for the Collection of Diagnostic Blood Specimens by Venipuncture; Approved Standard, Fifth Edition, H3-A5 Vol. 23 No. 32, December 2003.
3. NCCLS – Tubes and Additives for Blood Specimen Collection; Approved Standard-Fifth Edition, H1- A5 Vol. 23 No. 33, December 2003.
4. BD Evacuated Blood Collection System Package Insert 6/2004
5. NCCLS – Procedures for the Handling and Processing of Blood Specimens; Approved Standard-Third Edition, H18-A3 Vol. 24 No. 38, November 2004.
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