LABORATORY REAGENTS: CLASSIFICATION, PREPARATION AND FUNCTIONS - BIOLOGY

The laboratory reagents are chemical substances of various nature and composition that help the bioanalyst or professional clinician to conduct a series of reactions involved in the determination of a variety of clinical tests.

The reagents can be solid, liquid or gaseous according to their physical state, and according to their chemical nature they can be acids, bases, salts, alcohols, aldehydes, among others. All areas of a clinical laboratory require the use of various reagents, which differ from one another.

Bacteriology laboratory with a series of reagents. Source: Photo taken by the author MSc. Marielsa Gil.

The reagents can be purchased ready to use; some must be reconstituted, others must be prepared from the primary reagent mix. They can also be prepared with a simple dilution.

The function of each one will depend on the analysis to be carried out. For this, there are specific protocols that the analyst must follow carefully.

The handling and safeguarding of laboratory reagents must follow standards established by the institution, to guarantee the safety of the worker and their proper conservation. That is why it is recommended that reagents be stored according to the risk category established by government regulations.

Taking the risks into account and putting biosafety regulations into practice prevents workplace accidents.

Source: Prepared by the author MSc. Marielsa gil

Classification of reagents according to the degree of danger

There are at least 5 systems that classify reagents according to their degree of danger. These systems are those proposed by:

1) The United States Environmental Protection Agency (EPA).

2) The United States Department of Occupational Safety and Health (OSHA, Occupational Safety and Health Administration).

3) The European Community (EC).

4) The IMDG Code (International Maritime Dangerous Goods Code).

5) The United Nations (UN).

In general and coupled with what the EPA proposes, they can be classified as highly dangerous and dangerous.

-Highly dangerous reagents

They are those that can cause death to humans in very low amounts or that present a lethal dose in rats equal to LD ₅₀.

-Hazardous reagents

They are sub-classified as flammable, corrosive, explosive, and toxic.

Flammable substances

This category includes all reagents that contain more than 24% alcohol in solution or whose flash point is below 60 ° C.

Also included in this category are all materials that can easily cause fire by friction or by spontaneous chemical modifications or accumulation of their gases.

Likewise, those that are energetically incinerated when in contact with fire, and oxidizing reagents such as chlorates, nitrates, manganates and inorganic peroxides.

Toxic substances

Toxic reagents can be subdivided into irritants, anesthetics, asphyxiants, nephrotoxic, hepatotoxic, neurotoxic, and carcinogens.

Corrosive substances

Strong acids and bases fall into this category, that is, those that have a pH below 2.1 and above 12.4. These substances are so caustic that they can corrode steel.

Any trace of these reagents can react with other residues and form toxic compounds capable of endangering the integrity of workers.

These reagents should be away from the rest.

Reactive substances

They are reagents that react violently when combined with water, acids or bases, generating smoke, vapors or gases. This is how reactants that contain sulfur or cyanides in their composition react.

Explosive substances

They are substances capable of producing an explosion at 25 ° C at a pressure of 1.03 kg / cm ². This is because they have an explosive constant equal to or greater than that of dinitrobenzene.

Preparation of reagents

In general, the reagents should be prepared as follows:

Preparation of reagent from solid solutes

According to the technique, the precise grams are weighed on a scale. The solid solutes are placed in a beaker and a little of the solvent indicated by the technique is added, generally water is used. If necessary, the mixture is heated to aid dissolution, as long as the technique so indicates.

It should be allowed to cool before transferring to the volumetric flask. Use funnel for transfer. Wash the beaker with a little of the solvent being used and add it to the flask. Make up to the mark with the same diluent.

Transfer to a clean and dry bottle, label appropriately and store according to the reagent specifications.

Preparation of reagents from liquid solutes

The corresponding milliliters are measured with a serological or volumetric pipet. Do not pipet directly by mouth. Use propipette. Take care that the pipette does not fill with bubbles.

To do this, before sucking, make sure that the pipette is inserted to the bottom of the bottle and that it will continue to do so even after suctioning is complete.

Place the milliliters measured in a volumetric flask, make up to the mark with the diluent. The flask can be capped and inverted several times to mix. Transfer to a clean, dry container.

Filtration of reagents

Some reagents require filtration, for this purpose filter paper funnels are used. A smooth funnel is used if you want to recover the precipitate or a pleated funnel if the precipitate is not of interest.

Solid reagent transfer

A clean, dry spatula is used to take small amounts of solid reagents. And if the amount is a little larger, you can use a paper folded in the form of a channel to help slide the reagent to the other container.

Transfer of liquid reagents

In the transfer of highly corrosive liquids, spills and splashes should be avoided; For this, a funnel is used and a glass rod is placed through which the liquid to be transferred will slide.

If the reagent emits vapors, work under an extraction hood and use the necessary safety equipment (gloves, mouth cap or mask, safety glasses, gown). If the extraction hood is not available, work in a well-ventilated place.

Considerations after reagent preparation

The reagents after their preparation should be packaged in hermetically sealed containers, preferably amber.

Freshly prepared reagents must be carefully labeled with indelible ink, stating the name of the reagent, date of preparation, expiration date and the type of risk it presents (by inhalation, ingestion or contact).

Storage temperature is also important, the prepared reagent must be placed at the proper storage temperature. Some can be stored at room temperature, but others require refrigeration.

Safe handling of reagents

The reagents must be handled with care, preventing the inhalation of vapors, direct contact with skin or mucous membranes and their accidental ingestion. To do this, biosafety measures must be taken, such as the use of mouth covers, masks, gloves, safety glasses and a lab coat.

All these elements protect the person handling the reagents. Not all reagents emit vapors or are corrosive, so you have to learn to know them.

Before handling a reagent, check the bottle label and observe the safety pictograms; This will guide you on the preventive measures that should be taken. This will prevent possible accidents.

Scheme of old and current safety pictograms. Source: Lorenzo.profe

Reagents classified as flammable cannot be handled near a burner or burner in operation.

Reagent labels should always be placed on the bottle, never on the cap. The lids of the jars should not be interchanged, nor should they be placed on the table; they should be held with the fingers while handling the reagent.

Do not return the excess reagent taken to the original bottle, this can contaminate it.

If the reagent is corrosive or toxic you should never pipet by mouth, a propipette should always be used. As a safety measure in the bacteriology area, a piece of cotton should be placed on the pipettes on top, in case of accidents the cotton serves as a barrier.

When you want to dilute reagents such as strong acids, for example concentrated sulfuric acid or concentrated hydrochloric acid, it must be taken into account that water will never be placed directly on them, but the opposite; slowly the acid will be incorporated into the water, handling everything in a safety hood.

The work table must always be kept clean and dry. In case of spillage or fire, do not try to solve the incident with water.

Reagent storage

The bottle labels have a colored stripe that indicates which group it belongs to: flammable red stripe, corrosive white, reactive yellow, health risk blue, low risk green, white and red or white stripes, and incompatible black.

The items of the same group can generally go together and each group must be separated from each other. However, there are reagents that, even though they are from the same group, are incompatible with each other; they must be separated. Check the label for incompatibilities.

Acids and bases should never be stored together, likewise, flammable, corrosive, oxidizing reagents and peroxides should be as far apart as possible from each other (separate shelves).

Corrosive reagents should be at the bottom of the shelf and the most harmless ones at the top. Tall jars go to the top at the back of the shelf and small ones go to the front. Highly hazardous reagents require safety cabinets.

Finally, the MSDS (Material Safety Data Sheets) sheet for each reagent should always be read before handling.

Final disposal of chemical waste

The use of reagents generates waste materials that must be disposed of in such a way as to affect the environment as little as possible.

Most used reagents in a clinical laboratory, composition and function

Source: Table prepared by the author MSc. Marielsa gil

Source: Table prepared by the author Marielsa Gil