One must always remain alert and cautious while working in a laboratory. Careless handling of reagents, glassware or specimen to be tested in the laboratory can cause serious injury and can be dangerous to life. This chapter discusses hazards encountered in clinical biochemistry laboratory and measures to be taken to ensure safe working.
HAZARDS FROM DANGEROUS CHEMICALS
Toxic substances in a laboratory, can be absorbed either from direct contact through skin or by inhaling vapors or fine powder or can be swallowed by mistake while pipetting.
Injury results from the effect of these chemicals on other tissues such as bone marrow, liver and kidney.
Precautions for Avoidance of Accidents
- All bottles containing chemicals and reagents should be clearly labeled and the hazard noted.
- Never carry large bottles by the neck but hold the bottle with both hands.
- Keep bottle in use on shelves, not higher than eye level.
- Corrosive chemicals include strong acids or alkalis, e.g. nitric, sulphuric and hydrochloric acids, sodium and potassium hydroxide. Take great care while opening these bottles or when pouring from such bottles. Always add contents slowly to water, preferably while cooling and stirring.
- Never keep acids and alkalis in bottles with ground glass stoppers as they may get stuck.
- Whenever possible, use small measuring cylinders for measuring acids and alkalis. If more accurate measurement is required, use a pipette plugged with non-absorbent cotton wool or with a rubber tube attached.
- Toxic chemicals include cyanide and barbiturates. Keep these locked in a cupboard. Mouth pipetting for these should be totally forbidden.
- Organic solvents may have toxic properties. Thus, benzene is toxic to bone marrow. Carbon tetrachloride and other halogenated hydrocarbons are toxic to the liver. So keep exposure to the minimum. Carry out procedures including distillation in a fume cupboard or under a fumehood.
- Many chemicals have the potential to cause cancer and the most commonly carcinogenic chemicals used are aromatic amines such as benzidine and orthotolidine. Precautions include keeping them in well closed bottles labeled ‘carcinogenic’ and avoiding any contact with the skin.
- When handling carcinogens, rubber or plastic gloves should be used which must be washed well afterwards under cold running water.
- In case of accidental skin contact, wash in cold running water for several minutes.
- Explosion may be produced when using oxidizing agents, e.g. perchloric acid. Carry out work with perchloric acid in a fume cupboard.
- Ether kept in clear bottles exposed to sunlight can form peroxides. When redistilled, peroxides reach to sufficient concentration and it may produce a violent explosion. So keep ether in dark bottles.
- While distilling a solvent, the container must never be heated over a naked flame and efficient condensation of solvent vapor must be ensured all the time.
- Flammable gases stored in cylinders, constitute fire hazards, i.e. hydrogen, propane, acetylene. Keep cylinders, not in use, outside the laboratory.
- Bottles of chemicals and solutions should be handled carefully, and a cart should be used to transport a heavy or a multiple number of containers from one area to another.
- All bottles containing reagents must be properly labeled before adding the reagent.The label should bear the name and concentration of the reagent, the initials of the person who made up the reagent, the date on which the reagent was prepared, storage and potential hazards instructions (corrosive, toxic, irritants, flammable, explosive).
- Disposal of flammable solvents in sanitary sewers is not allowed.
- Acids, caustic materials, and strong oxidizing agents should be mixed in the sink. Where water is available for cooling.
- Do not throw any solid into the sink. If you have to pour strong acids or alkalis make sure that you let a lot of tap water rinse it away.
- All the electrical equipments should be grounded using three-point plugs and use of the extension cord should be prohibited.
- Every laboratory should have the necessary equipments to put out a fire in the laboratory. All staff should be familiar with the location of fuse boxes and circuit breakers. Any electrical sparking should be reported immediately.
INFECTION HAZARD
Infection hazards are mainly viral hepatitis and acquired immunodeficiency syndrome (AIDS). The guidelines for the laboratory handling of material from patients with AIDS and viral hepatitis are similar.
AIDS: It is caused by human immunodeficiency virus (HIV) which by damaging T-cells, reduces the ability of an affected individual to resist other infections. The virus has been detected in blood, breast milk, semen, vaginal fluid, saliva, tears, urine and brain tissue. In the laboratory, the infection is most likely to occur by infection of exposed cuts and by accidental self-inoculation with blood containing virus.
Viral hepatitis: It occurs in three forms. Hepatitis A has an incubation period of 14 to 35 days. It occurs in general population. Urine and feces contain the virus. Diagnosis requires demonstration of the virus in the feces or presence of hepatitis specific IgM in the serum. In the laboratory, entry of virus is usually through ingestion of contaminated food or drink or by placing contaminated fingers or other objects in the mouth.
The second form, hepatitis B, has a longer incubation period of 40 to 120 days. During the acute phase of illness, blood is highly infectious but urine and feces are probably infected. Third form is non A-non B hepatitis. The virus occurs in blood. Both hepatitis B and non A-non B hepatitis are associated with a carrier-state, so blood may continue to be infectious long after the patient is well. Blood is the main source of infection in the laboratory. Possible routes of entry are through direct entry due to breaching of skin surface or through ingestion.
Precautions
- Precautions include special handling of high risk samples. But this does not exclude the possibility that other specimens are not infected. Remember that even blood from a healthy person may be dangerous. It may contain microorganisms that can 3cause jaundice. So be just careful with blood specimen as you are with specimen of stool and CSF. So it is wise to treat all samples as potentially infectious samples. So mouth pipetting for all the samples should be avoided completely. Try not to get blood on your hand and do not spill blood in the laboratory.The AIDS virus can spread through the use of syringes, needles and instruments, which have been in contact with the blood of a person who is carrying the AIDS virus, even if he is not sick. It is, therefore, important to only use sterile syringes, needles and instruments. The virus is very fragile and dies at only 56°C or when soaked in common disinfectant.There are three ways to sterilize equipments:
- Completely immersing instruments in boiling water for 10 minutes.
- Soaking for 20 minutes in disinfectant solution:
- Household bleach—1% solution (available in market as 3.5% readymade solution polar R Dilute it with 2.5 volumes of tap water to give 1% solution
- Sodium hypochlorite—1% solution
- Glutaraldehyde—2% (available commercially as Cide)
- Lysol—2.5% solution
- Hydrogen peroxide—3% solution
- Dettol—4% solution
- Savlon—5% solutionA solution of sodium hypochlorite prepared daily is inexpensive, effective and easily available disinfectant. Also concentration ranging from 0.1 to 2% of chlorine is effective depending on the amount of concentration. All the linen soiled with blood/body fluid should be dipped in 1% household bleach or sodium hypochlorite for half-an-hour washing.Instruments that require sterilization or disinfection should be thoroughly cleaned before being exposed to disinfectants.
- Sterilization and disinfection: Sterilization is a process of freeing an article from all living organisms including bacterial, fungal, spores and viruses. Moist heat (steam) under pressure using different type of autoclaves is the most dependable method of sterilization. Specially modified pressure-cookers at 121°C at 5 lbs/sq inch pressure for 15 to 20 minutes gives high level of sterilization.
- Personal protective equipment (PPE) is the primary barrier of protection worn before handling potentially hazardous biological materials. PPE include laboratory coats and gloves. In case of any cut on hands, cover them well with plastic tape and then wear gloves. Laboratory coats should not be kept in the lockers used for personal clothes. While wearing gloves avoid touching general items like telephone, keyboard, etc. When leaving the laboratory, protective clothing should be removed.
- Proper methods of self decontamination after performing work are to be followed, i.e. to remove the laboratory coat first, then gloves, and then thoroughly washing the hands with soap or disinfectant. If hands are washed before removing the coat, the hands may become contaminated again while handling the coat.
- The following are few safety precautions to be followed when pipetting in the laboratory:
- Do not draw reagents or specimens through pipettes directly by mouth. Use automated pipette.
- Pipette contents should be allowed to run down along the wall of the container, making sure not to release the contents from a height.
- Avoid performing mouth pipetting and never blow out pipettes that contain potentially infectious material, e.g. serum.
- Do not mix potentially infectious material by bubbling air through the liquid, which leads to aerosol formation.
- Put needles and sharps in puncture resistant containers.
- Staff locker rooms with washing facilities should be separate from the working area.
- Do not eat, drink or smoke in the laboratory and never store the food or drink in the refrigerator. An entirely separate staff room must be made available to enable food to be kept out of the working area.
- Periodically clean out freezer and dry ice chests to remove broken ampules and tubes of biological samples. Use rubber gloves during this cleaning.
- Hands should be washed as soon as possible when they come in contact with potentially infectious materials. A vigorous handwashing with an approved agent is appropriate. Hands must always be washed before leaving the laboratory, and immediately if visible contamination occurs.
FIRST-AID AND EMERGENCY TREATMENT IN THE LABORATORY
- Acid burns: Nitric, sulphuric, hydrochloric and trichloroacetic acids.
- For acid splashes on skin, first wash thoroughly and repeatedly with water and then bathe the affected skin with cotton wool soaked in 5% aqueous sodium carbonate.
- In case of acid splashes on the eye, wash the eye immediately with large quantities of water, sprayed from a wash bottle or a rubber bulb or hold the eye under the running tap. After washing, put 4 drops of 2% aqueous sodium bicarbonate into the eye. Continue to apply bicarbonate solution in the eye until checked by the doctor.
- In case of accidental swallowing of acids while using a pipette, call a physician. Make the patient drink some 5% soap solution immediately. Alternatively, give him two whites of egg mixed with 500 ml of water or milk. If neither is available, give ordinary water to drink. Make him gargle with the soap solution. If the lips and tongue are burned by acid, first rinse thoroughly with water and then with 2% aqueous sodium bicarbonate.
- Alkali burns: Sodium, potassium and ammonium hydroxide.Alkali burns are as serious as, and often more serious than acid burns.
- In case of alkali splashes on the skin, wash thoroughly and repeatedly with water and then bathe the affected skin with cotton soaked in 5% acetic acid (or undiluted vinegar).
- For alkali splashes on the eye wash immediately with large quantities of water sprayed from a wash bottle or rubber bulb. Squirt the water into the corner of the eye, near the nose. After washing with water repeatedly, wash the eye with a saturated solution of boric acid.
- In case of accidental swallowing of alkalis while using a pipette, make the patient drink at once 5% solution of acetic acid or lemon juice or dilute vinegar (1 part vinegar to 3 parts water). Make him gargle with the same acid solution. Give him 3 or 4 glasses of ordinary water. If the lips and tongue are burned by the alkali, rinse thoroughly with water and bathe with 5% acetic acid solution.
- Burns caused by heat: They fall into two categories:
- Severe burns affecting large area of skin, e.g. burns caused when burning ether or boiling water is spilled over victim.
- Minor burns affecting a small area of skin, e.g. burns caused by hot glassware or a Bunsen flame.
Severe burns: In this case, if the victim is on fire, e.g. if splashed with burning ether or other inflammable solvent, roll him in a blanket or overall to smother the flames. Do not remove his clothing. Do not apply any treatment to the burns. This must be left to the physician.5Minor burns: Plunge the affected part into cold water or ice water to soothe the pain. Apply mercurochrome or acriflavine ointment to the burns. Apply a dry gauze dressing loosely. Never tear off the blisters that form over the burns. - Poisoning: This can be caused by:
- Inhaling toxic vapors or gases (e.g. chloroform).
- Accidental swallowing while pipetting a poisonous solution. In all cases, place the victim in the open air while waiting for the physician.
- Injuries caused by broken glass: These are caused by broken test-tubes, syringes or other glassware. So wash the wound immediately to remove any glass piece and apply acriflavine ointment to the wound. Cover with gauze and adhesive tape. If the cut bleeds heavily, stop the bleeding by pressing down on it with a compressor. Refer the patient to a physician.
- Contamination by infected material: For wounds caused by broken glassware containing stools, pus, etc. wash the wound immediately. Check whether the cut is bleeding. If not, squeeze hard to make it bleed for several minutes. Bathe the whole area, i.e. the edges of the cut and inside the cut, with antiseptic lotion. Wash thoroughly with soapy water. Bathe again with antiseptic lotion. If the material involved is known to be very infective, e.g. pus, refer the patient to the physician.If infected material is accidently sucked into the mouth, spit it out immediately and go in for forced vomiting. If skin is infected by highly virulent organism touch the involved part with pure carbolic acid.
CASE REPORTS
- In the blood collection room of emergency, while withdrawing blood sample there was spillage of blood on the table. How should the table be cleaned?Universal precautions about HIV prevention are required to be taken in every patient walking in side the emergency room. So the blood should be soaked with the absorbent swab material soaked in 10% bleach solution. It should be kept for 20 minutes and then disposed off.
- How is infective waste disposed off in health case setting?The infected waste in health case setting can be disposed off by either incineration or deep burial with bleach or lime. Liquid wastes, blood, body fluids placenta, etc. should be buried in deep pit by covering with bleaching power or lime. Solid waste, dressing, laboratory and pathology wastes should be incinerated.
- What steps to be taken on exposure to HIV infected blood/body fluids and contaminated sharps?The following steps should be taken:
- Needle sticks and cuts should be washed with soap and water
- Splashes to the nose, mouth or skin should be flushed with water
- Eyes should be irrigated with clean water, saline or sterile irritants
- Pricked fingers should not be put into mouth reflexively.
No scientific evidence exists as to the fact that the use of antiseptic for wound care of squeezing the wound will reduce the risk of HIV transmission. But this must always be done.Exposure should be reported promptly to the appropriate authority because in some cases, HIV, post-exposure prophylaxis (PEP) may be recommended and treatment started as soon as possible. Success of PEP therapy is maximal when started within a matter of hours after the exposure. The decision to start PEP is made on the basis of degree of exposure to HIV and HIV status of the source from whom exposure/infection has occurred. Patient doubtful of having HIV infection should be referred to voluntary counseling and testing center (VCTC). - What are the NACO guidelines for post- exposure prophylaxis?Post-exposure prophylaxis (PEP) is any preventive medical treatment started immediately after exposure to a pathogen, in 6order to prevent infection by the pathogen and the development of disease.This includes:
- First aid
- Counseling
- Risk assessment
- Relevant laboratory investigations based on informed consent of the source and exposed person
- Depending on the risk assessment, the provision of short term (4 weeks) of antiretroviral drugs
- Follow-up and support.
- How sodium hypochlorite solution is used for cleaning of blood/body fluids/other potentially infectious material spills.Work surfaces are to be decontaminated at least twice a day and after every spill of viable/infected material. Sodium hypochlorite available as 4% w/v solution is used as follows:
- Blood spillage is cleaned up using 1% hypochlorite solution. (Mix 1 part of readymade 4% w/v sodium hypochlorite solution with 3 parts of sterile water.)
- Minor contamination of floor, furniture, fittings and equipments must be cleaned with 0.1% hypochlorite solution. (Mix 1 part of the prepared 1% hypochlorite solution and mix with 9 parts of sterile water). Procedure of cleaning up blood/ body fluid spills using sodium hypochlorite solutionProcedure of cleaning up blood /body fluid spill.
- Alert the people in vicinity immediately about the spillage.
- Put on appropriate safety equipment.
- Carefully pour freshly prepared 1% sodium hypochlorite solution around edges of spill and then into the spill.
- Avoid splashing.
- Allow a 20 minutes contact period.
- Use absorbent tissue/any other material to wipe up the spill, working from periphery towards center.
Then, dispose off the tissue after keeping it dipped in 4% hypochlorite or 10% bleach.
- Why a fresh solution of hypochlorite should be used?According to the literature, depending on storage conditions; all hypochlorite solutions lose half of their potency in less than thirty days. The biggest factors for this are light, temperature and the time, since it is manufactured.The biggest misconception is that liquid household bleach (sodium hypochlorite) does not lose potency until a sodium hypochlorite solution is made. But the fact is that it is already a sodium hypochlorite solution, that starts degrading soon after manufacture, so a bottle bought from the market is not a fresh hypochlorite solution, it is a hypochlorite solution with an unknown chlorine content. When we make a solution all we are doing is diluting an already weak hypochlorite solution even more.For a chlorine solution to be a good disinfectant it must meet the chlorine demand. The chlorine demand is the amount of free available chlorine, or (FAC) often called hypochlorus acid or HOCl, needed to disinfect or oxidize organic matter. If the chlorine demand is not met then complete disinfection has not been obtained. One of the best signs that the chlorine demand has not been met is the strong chlorine odor present produced by chloramine gas. HOCl, is the most germicidal of all chlorine compounds, between 80 and 120 times stronger than the OCl− ion. The pH of the solution is the determining factor on which species of chlorine is produced in the solution, HOCl or OCl− ion. The ideal pH of a disinfecting chlorine solution is a pH of 6 to 8.
- What are the steps for handling of chemical spills?The following steps are to be followed:
- Immediately vacate the room if the spill represents a hazard which is immediately dangerous to life and health.
- Contact occupational safety and health services emergency spill response coordinator.
- Area of spill to be secured from personnel entering the danger zone.
- Be aware of the nature of spill, it may be helpful in further management.