Preparation Of Standard Solutions Biology Essay

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The analytical balance is one of the accurate and most basic instruments used in the clinic laboratory. It is a high precision instrument which is used for the preparation of standard solutions and wherever accuracy of 1 mg or even lesser. Analytical balances mostly have a capacity of 200g. Semi microbalances are capable of weighing up to 100g and microbalances have a capacity of 20 to 50g. A typical analytical balance would have a lever with two equal arms and a pan suspended from each arm which is balanced by each other.

The mass to be weighed is placed in one pan, and a standard mass in the other pan until the beam is as close to equilibrium as possible. In more accurate balances, a slider mass is moved along a graduated scale. The position of the slider gives a fine correction to the mass value. They are mainly used for weighing chemicals in laboratories.

Now a days we do have modern electronic balances, whereby the weight of the object is easily found by placing it on the pan. It is connected to a digitalizer which gives the accurate weight of the object.

chemical_balance

Types of analytical balances:

There are three types of analytical balances:

Double pan balance

Single pan balance

Electronic balance

The two main properties of an analytical balance are accuracy and precision.

Accuracy is the ability of the measurement to match the actual value. In other words, it is the degree of closeness to the actual value.

For E.g. If the actual weight of the object is 2.545 g, if the measured value is 2.542 then it is said to be accurate.

Precision is the reproducibility of the same value on each an every measurement. As more and more trials are made, the precision improves and the mean value generally becomes a better and better estimate of the true value.

Double beam balance:

It consists of a beam supported at the centre by a knife edge made of steel or agate that rests on a plate of hard material which is fixed at the top of a support that moves inside the pillar. On the two ends of the beam are two knifing edges pointing in the upward direction and equidistant from the center. Two stirrups that carry scale pans of equal masses are hung from them. To the middle of the beam a pointer is attached that is perpendicular to the pans and it moves over an ivory scale fixed at the bottom of the pillar. When the balance is not being used the beam is kept in fixed position by turning a handle. The complete arrangement is enclosed in a wooden box with glass doors which are kept closed during the time of weighing as well as when not using the balance.

Physical Balance_b

Single Pan Balance:

More accurate weights can be measured using a single pan balance. The weight is measured in single pan balance by way of one or more bars (levers) with sliding weights which are marked with the exact weight that balances the contents in the pan. The principle of operation of single pan balance is a simple lever with weights on one side of the fulcrum which balances with the weight of the unknown object in the pan.

MetH10

Electronic balance:

The latest analytical balances are electronic balances which have can measures masses to the range of 0.0001 g. The weighing pan is of high precision ranging from 0.001 mg to 100 g. They are kept inside a glass hood with doors in order to keep it free from dust as it is so sensitive and also to prevent any air currents in the room affect the balance's operation. There are a lot of sophisticated balances available in today's world for accurate measurement of the weight.

Electronic-Balance-ES-500HA-

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3.3. PRINCIPLE

A balance determines the weight of the object by comparing the mass of two sets of objects wherein one pan has an unknown mass and the other pan has the computable weights. At the time when both the pans are in equilibrium and the fulcrum is not tilted towards any side then the balance is supposed to give the accurate weight of the same.

The principle for single pan balance is that the weight is measured by means of one or more bars (levers) with sliding weights that are marked with the exact weight that balances the contents in the pan. So the principle is a simple lever with weights on one side of the fulcrum that balanceS the weight of the unknown object in the pan.

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3.3. WORKING & MAINTENANCE

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Working:

The main things to be remembered while weighed in an analytical balance are:

Balance environment

Calibration of the balance

Errors during weighing

Balance environment:

The balance should be placed on a smooth surface

The location should have low amount of disturbance and air current

The place surrounding the balance should be neat

Constant power supply in case of single pan balance and electronic balance is necessary

Calibration of the balance:

The power is turned on 15 minutes before weighing so as the balance starts calibrating.

The leveling screws should be adjusted in order to ensure that the plummet comes precisely over the index so that the pillar may become vertical.

Once the balance is calibrated it should not be disturbed by moving it or so, because once it is moved it loses its calibration

Errors during weighing:

There are two kinds of errors.

Instrumental errors

Personal errors

Instrumental errors:

These are errors which are in the balance itself such as the pans are not in proper position, no proper balancing, etc. This can be corrected by proper calibration of the instrument.

Personal errors:

These kind of errors are due to inexperience of the observer such a taking the reading without proper setting of the apparatus, taking readings without proper precautions, etc.

How to weigh in an analytical balance:

First keep all the necessary weights and the chemical to be weighed, spatula, hair brush, butter paper, etc.

The object to be weighed must be at the room temperature and should not be either too hot or too cold.

Utilizing the hair brush, the pans should be cleaned gently.

Make sure that the instrument is working properly by turning the handle release of the beam which should be resting on its knife edge. The pointer moves within the ivory scale. The handle is turned back to its normal position.

Right pan should be used for the objects whose weight is to be determined and the left pan must be utilized for the standard masses from the weight box.

The weights must be added in descending order of their value which means the heavier weight should be put in first and then the lower ones. The weights must be added one by one.

The correct weight to measure is kept on the left side of the pan.

Start adding the chemical on the right pan. Make sure that the pointer is in resting position.

Now the handle of the beam is turned. Once turned the pointer starts moving over the scale.

When it moves towards right the weight is more, when it moves towards the left the chemical is more.

The correct weight is indicated by the pointer which comes to the center .

The handle is turned back to its original position.

Care should be taken that the doors of the balance are closed when the pointer moves, as the air current will cause fluctuations.

Once the weighing procedure is complete the object and weights must be removed from the pan and the balance should be checked as whether or not it is properly clean.

Care & maintenance of Analytical Balance:

The balance must always be kept in a glass case to keep it free from dust & moisture.

Inspect the balance for cleanliness. If necessary, using a hair brush, dust the pans, etc., taking special care not to jar the balance.

Determine zero point before "adjusting the zero." Keep a record of the zero point before adjustments are made, so that you may have an indication of the general constancy of the balance.

Liquids should be weighed in a closed container so that none of the contents is lost.

Move rider, chain, or other device equivalent to adding 1.0 or 2.0 mg. on the right pan so as to determine whether the beam swings freely on release.

If the balance is not damped, let the beam swing 3 or 4 times over about half of the length of the index scale. Note the uniformity of "falling off," or whether there is any overshooting.

If the balance is damped, release and let it come apparently to rest. Note readings 3 or 4 times about 5 seconds apart. Watch for drift or any irregular change in reading, which might indicate any unsatisfactory weighing conditions.

Re-determine zero point as a check on the constancy of the balance.

If anything is spilled on the pan then take the pan out and clean it thoroughly.

The balance should not be loaded with a weight greater than what it is constructed for.

Weights should not be touched by hand and must be lifter with forceps as the dirt or grease may stick on to the weight changing its accuracy and in turn affecting the precision of the instrument.

After use close balance case tightly, and see that all arrestments are in proper position.

If there is a special dust cover, outer case or other extra protection, then ensure that it is used.

REAGENTS

A reagent is a substance that when added to a chemical reaction brings about chemical change or to see if a reaction takes place. It is used to detect measure, examine, or make other substances.

For e.g., iodine when added to a solution, when gives violet color indicates the presence of starch. Thus iodine is a chemical reagent which brings a chemical change in the starch solution.

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3.4. PREPARATION OF REAGENTS

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Methods for Preparing a Reagent:

One of the most important requirements for the successful completion of any protocol is to prepare reagents accurately.

The general guidelines for the preparation of reagents are

• Use of clean glass- and plastic ware,

• Use of high-purity reagents,

• Safe handling of chemicals and biochemicals

• Use of high-purity distilled or deionized water,

• Accurate weighing,

• Pipetting and pH measurement

• Avoidance of chemical or microbial contamination of reagents.

Use of clean glass- and plastic ware:

The glass wares which we use should be thoroughly washed with tap water and then rinsed with distilled water so as to prevent any contamination of the previous reagent.

Use of high-purity reagents:

The reagents which are used should be of good condition. Old or expired chemicals will not give a proper result. The purity of the chemicals is also important and thus the chemicals are immediately closed once it is weighed.

Safe handling of chemicals and biochemicals:

The chemicals should be handled with care as certain chemicals will be carcinogenic (cancer causing), others being corrosive (E.g. HCl), etc. Hands should be thoroughly washed after preparing the reagents. Wearing a lab coat while preparing reagents is compulsory.

Use of high-purity distilled or deionized water:

Water also plays an important role in preparation of regents. Distilled water alone should be used for preparing reagents. For fine reagents double distilled water can also be used.

Accurate weighing

Weighing accurate quantity of chemicals is important as less or more amounts of the chemical will hinder the activity of reagent. If the chemical reaction is a quantitative analysis, then weighing should be accurate for better results.

Pipetting and pH measurement:

Pipetting errors will also affect the activity of reagent.

For e.g. if 2 ml of the reagent is added instead of 5 ml then the reaction will not have the sufficient reagent for the reaction to take place and results will be wrong.

In case of biochemical reactions, pH plays an important role as the reagent will work only if the pH is correct. The pH is adjusted using a pH meter.

Avoidance of chemical or microbial contamination of reagents:

Once the reagent is prepared it is stored in a proper place in a suitable environment such as in the shelf or in the refrigerator. Some reagents should be stored only in refrigerator so as to prevent contamination. Once contaminated the reagent cannot be used further. So, proper maintenance of the reagent is necessary.

It is important that the reagents prepared in the laboratory have effective quality. In addition, it is vital to document particular storage conditions and expiration dates, as suitable.

The methods utilized by laboratories may vary with respect to the accurate concentrations and/or volumes of reagents utilized, reagent container labeling procedures, and documentation.

Being careful and specific is important for preparation of reagents.

Common Reagents Used in Biochemistry

Acetic Acid which is an organic compound gives vinegar a sour taste and also a strong odor. CH3COOH is the chemical formula for acetic acid. It is corrosive and its vapor can be harmful for the eyes, nose, and throat. It is an important chemical reagent which is commonly utilized in industrial chemistry in the production of polyethylene terephthalate which is mainly utilized in soft drink bottles, cellulose acetate used mainly for photographic film and polyvinyl acetate for wood glue, as well as synthetic fibres and fabrics. With a worldwide requirement for acetic acid at 6.5 million tonnes per year it is a quite vital reagent.

Ampicillin falls in the penicillin group of beta-lactam antibiotic which has been utilized to treat bacterial infections. Ampicillin is able to penetrate gram-positive and some gram-negative bacteria, however there are strains with ampicillin resistance genes. It is considered part of the aminopenicillin family. C16H19N3O4S is its chemical formula. From a rash to potentially lethal anaphylaxis varies the allergic reactions to ampicillin. This has been in use widely from 1961 onwards and is being utilized even presently.

Cobalt Chloride is a chemical utilized to formulate reagents. It is a small, blue colored crystal which is odorless and stable under normal atmospheric conditions. The term cobalt chloride (CoCl2) has also been used to describe it's hexahydrate form which turns its typical blue into a deep rose-red color (CoCl2:6H20) making it a good indicator for water. It is used mainly in inks, varnishes, enamels, fertilizers, feed additives and humidity indicators. But the role in reagent formulation played by this chemical is different from the uses mentioned here above.

Ammonium Persulphate, APS,[(NH4)2S2O8] is designated electrophoresis grade and is utilized largely in biochemistry and molecular biology laboratories. For making the polyacrylamide gels APS is utilized. It is known as an extremely oxidizing agent, and is combined with tetramethylethylenediamine to aid in the polymerization process by catalyzation and forming polyacrylamide gel. However, APS should be handled with care as it is a hazardous material. Exposure to the chemical requires proper protection in the way of gloves and eye protection. For the preparation of dyes and photography this chemical is used, as well as when dissolved in sulfuric acid, it is used as a cleaning agent for laboratory glassware.

Tollens Reagent is frequently utilized in the finding of aldehydes. These same aldehydes are then oxidized to carboxylic acids. In case the Tollen's reagent detects an aldehyde, then the test results in a silver mirrored solution. On the other hand if there is no aldehyde present then a yellow mirrored solution may result. Tollen's reagent is also used to reduce Silver (I) to silver metal. In Organic Chemistry Labs Tollen's reagent is frequently utilized by students in their experiments.

Glycerol is used in both agarose gel electrophoresis and polyacrylamide gel electrophoresis. Glycerol when added to a sample of DNA or protein increases the density of that particular sample, resulting in the sample to fall down into the gel way instead of floating away to mix with other samples of the loading buffer. In brief, adding glycerol helps assure that samples remain in the gel during the experiment which is of prime importance to the receipt of accurate results in experiments involving gel electrophoresis.

Sodium Dodecyl Sulfate is another common reagent utilized in the preparation of proteins for polyacrylamide gel electrophoresis. SDS is a detergent (soap) that can dissolve hydrophobic molecules, but also has a negative charge (the sulfate) attached to it. SDS denatures proteins by disrupting their non-covalent bonds. SDS produces a negative charge on the protein treated with it. Interestingly, the reagent is also used in toothpaste, shampoo, and shaving cream preparation.

Ethidium Bromide is a chemical that is often mixed with agarose gels and used to visualize DNA. In a process called intercalation, ethidium bromide stacks itself between the base pairs in DNA. Once there, the ethidium bromide becomes excited, and fluoresces a bright pink color when exposed to ultraviolet light. This is due to the hydrophobic interactions that are inside the double helix of DNA. This bright pink color in turn allows researchers to detect the position of DNA on the agarose gel. Although it is useful, ethidium bromide should be handled with care and proper lab safety techniques. This is because it is a known mutagen. Therefore, you should not have any skin to skin contact with this chemical, including consumption. Using gloves with this chemical to avoid it touching your hands is advised. It would also be wise to use eye protection when handling this chemical.

Dragnedroff's Reagent is a reagent consisting of a mixture of bismuth subnitrate and potassium iodide dissolved in aqueous acetic acid. This test is used to detect alkaloids and amines/organic bases. When it tests positive for amines, it produces a visible color and can be used for comparison purposes.

IPTG, or Isopropyl β-D-1-thiogalactopyranoside, is a common reagent used in the biochemical over-expression of a target protein in an organism. This is where the DNA sequence of the protein can be over made, or cloned into smaller plasmids. IPTG is used in the induction of protein over-expression, typically in conjunction with the lac operon. It is not considered a health hazard and does not require any special handling or treatment.

Methyl Orange is an indicator dye that is synthesized through a reaction between sulfanilic acid dissolved in dilute acetic acid, sodium nitrite and dimethylaniline. In the presence of acidic conditions, methyl orange changes from a red to an orange color.

Cleland's Reagent is commonly used in the protection of sulfhydryl groups in cells. It readily permeates cell membranes and thus allows enzyme activity to be restored due to the in vitro oxidation of the sulfhydryl groups. Cleland's reagent is also known as (D,L)-1,4-Dithiothreitol, or DTT. DTT is an unusually strong reducing agent, owing to its high conformational tendency to form a six-membered ring with an internal disulfide bond. The most common use of DTT is as a reducing or "deprotecting" agent for thiolated DNA. DTT is usually mixed with a DNA solution and allowed to react. The DTT is then removed from the mixture by filtration or chromatography in a process known as "desalting." DTT is also commonly used to reduce the disulfide bonds of proteins.

Biuret Reagent is used in biochemistry as a protein assay. The biuret compound is equivalent to two molecules of urea, without one of the ammonia groups. This assay is useful when protein yield is not a problem. The assay is run, and interpreted using a UV spectrophotometer with the absorbance reading at 550nm.

Bradford Reagent is used in protein assays. It is used to determine the concentration of a protein in solution. In the presence of a protein, the reagent turns blue and in acidic conditions, the reagent turns red. It contains 5% ethanol and 9% phosphoric acid along with a small amount of Coomassie Blue stain and some sodium lactate. It is a corrosive substance and requires the use of gloves and safety glasses.

BCA is another reagent used in protein assay, like the Bradford Reagent. However, BCA does not interact with many substances other than proteins, giving it an advantage over many other reagents. The BCA reagent forms a complex with copper, giving it a strong absorbance at 562nm for results.

Sephadex G-50 is a cross-linked dextran gel used frequently in gel permeation chromatography. It is manufactured in bead form and can be used in conjunction with ammonium bicarbonate as an elution agent in gel chromatography.

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3.5. FORMULATION & PREPARATION OF REAGENTS

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There are two ways of preparing reagents

• Dilution of reagent

• Using solid chemicals

Dilution of reagent:

This method employs the conversion of the stock reagent to the working concentration using distilled water.

In order to properly dilute a reagent, one must calculate the correct and proper volume of the reagent needed to yield a desired concentration.

For e.g. if 10% sodium chloride solution is given and if you need only 5% solution then dilution is done by doubling the volume of the reagent using distilled water.

Using solid chemicals:

Here the chemicals are weighed in the chemical balance and then dissolved.

For e.g. dissolving 5g of sodium chloride in 100 ml of distilled water gives a 5% solution of sodium chloride.

The preparation of few of the reagents used in the medical lab is as given under -

Acetic Acid 10% (100gm/lit) -

Formulated by mixing 20 ml Glacial acetic acid (CH3COOH) with distilled water qs (quantity sufficient) 200 ml.

Acid Ethanol -

It is required for modified Ziehl Neelsen Stain. Prepared from 3 ml Hydrochloric Acid and 70% Ethanol upto 100ml.

Barium Chloride 10% (100gm/lit) Aqueous Solution -

Formed by 10 gm Barium Chloride (BaCl2) and distilled water qs 100 ml.

Benedict Qualitative Solution -

The required ingredients are 173.3 gm Copper sulphate (CuSO4.5H2O), 173 gm Trisodium citrate (Na3C6H5O7.2H2O), 100 gm of Sodium carbonate (Na2Co3) anhydrous and 1000 ml of Distilled water.

The copper sulphate is dissolved by heat in 100 ml of water. Simultaneously Trisodium citrate and sodium carbonate is dissolved in 800 ml distilled water. After this slowly add the copper sulphate solution to the trisodium citrate & sodium carbonate solution, stirring constantly across the addition process. Now make up the total volume of the solution to 1000 ml by adding qs distilled water.

Buffered Water -

This is needed for Giemsa and Leishman stains. The ingredients required are 3.76 gm of sodium hydrogen phosphate (Na2HPO4.2H2O.), 2.10 gm of Potassium dihydrogen phosphate (KH2PO4) anhydrous and qs 1000 ml distilled water. By making use of narrow range pH paper adjust the pH of solution to keep it in the range of pH7.0 to pH7.2.

Buffered Water -

This one is for JSB stain and prepare by mixing 0.417 gm of Disodium hydrogen phosphate, 0.752 gm of Potassium acid phosphate and 2000 ml of distilled water. The pH to be maintained for this solution is in the range of 6.2 to 6.8.

Carbol Fuchsin -

It is required for Ziehl Neelsen stain. It is made up of two solutions A & B. Their preparation is as under -

Solution A - saturated solution of basic fuchsin - prepared by mixing 3gms of basic fuchsin and 100 ml of 95% ethanol solution.

Solution B - phenol solution 50gm/lit (5%) aq - prepared by mixing 10gms phenol and 200ml distilled water.

Now after preparation of both the solutions, mix the two solutions in volumes of 10ml of solution A and 90 ml of solution B.

Carbol Fuchsin -

This is needed for modified Ziehl Neelsen stain and obtained by mixig 5gm basic fuchsin, 10ml 95% ethanol and 90ml 5% phenol solution.

Cary-Blair Transport (Holding) medium -

It is prepared by mixing 1.5gm sodium thioglycolate, 1.1gm disodium hydrogen phosphate anhydrous, 5gm sodium chloride, 5gm agar and 991ml distilled water. The solution must be prepared in chemically clean glassware and the contents during mixing must be heated till the time the solution becomes clear. Once the solution cools down to a temperature of 500C the add 9ml of freshly prepared aq calcium chloride 1%. Now adjust the pH to around 8.4. After this pour 7ml in thoroughly rinsed and sterilized 9ml screw capped vials. Then steam the vials containing media for 15 minutes and after that cool and tighten the cups.

Crystal Violet -

This is made by mixing two solutions 1 & 2.

Solution 1 - prepared by mixing 2gm crystal violet with 20 ml 95% ethanol solution.

Solution 2 - prepared by mixing 0.8gm ammonium oxalate and 80ml distilled water.

Once both the solutions are ready then mix the two and store for one day before utilizing the same. After that filter through a filter paper into a staining bottle.

Dichromate cleaning solution -

This solution is used for cleaning glassware and obtained by mixing 100gm potassium dichromate in 1000ml distilled water. After that 100ml pure sulphuric acid is added to the dichromate solution. While adding the acid it must be done carefully and slowly and ensuring that acid is added to water.

EDTA - Dipotassium salt solution (10%)

It is prepared from 20gm dipotassium ethylene diamine tetra-acetate and qs 200ml distilled water.

Eosin (2%) solution in Saline -

Obtained from mixing 3gm eosin and qs 100ml NaCl(0.85%) in aqueous solution.

Fouchet Reagent -

Before preparing the reagent we need to first prepare 10% solution of ferric chloride which is obtained by mixing 10gm Ferric chloride and qs 100ml distilled water. Once the ferric chloride solution is ready then take 10ml of trichloroacetic acid and dissolve in 70ml distilled water in a volumetric flask. After that add 10ml of ferric chloride solution and then make the volume 100ml with qs distilled water.

Giemsa Stain -

The ingredients for this are 0.75gm powdered Giemsa stain, 65ml methanol and 35ml glycerol. Transfer all the constituents in a bottle containing glass beads and then shake the mixture for 3 times a day for 4 consecutive days. After that filter the solution and use.

Gram Iodine solution -

The required ingredients are 1gm iodine, 2gm potassium iodide and 100ml distilled water. Firstly mix potassium iodide in 30ml water. Then add iodine and mix until dissolved. After that add 70ml water and mix well and then store in a brown bottle.

Hydrochloric Acid (0.1N) 0.1mol/lit -

It is obtained from 8.6ml concentrated HCL and qs 1000ml distilled water. Take 500ml water and add acid drop by drop. After that add water till the volume becomes 1ltr. The solution must be utilized within 1 month of preparation.

Jaswant Singh and Bhattacharya Stain (JSB)

There are two JSB solutions, JSB solution 1 & 2.

JSB Solution 1 - The ingredients required are 0.5gm methylene blue, 3ml 1% sulphuric acid, 0.5gm potassium dichromate, 3.5gm disodium hydrogen phosphate dihydrate and 500ml distilled water. First of all dissolve methylene blue in distilled water. Then slowly pour sulphuric acid, adding 1ml at a time and stirring & mixing it continuously. After that add potassium dichromate, a purple precipitate is formed. The add disodium hydrogen phosphate dehydrate. On stirring it for some time the precipitates get dissolved. After this boil the solution in a flask for about 60 minutes along with a reflux condenser till the blue colour of the solution deepens.

JSB solution 2 - Obtained by mixing 1gm eosin yellow which is zinc free with 500ml distilled water.

Leishman Stain -

The ingredients required are 1.5gm Leishman powder and qs 1000ml methanol. Firstly rinse out a clean bottle with methanol and add a few clean dry glass beads to it. After that add the staining powder followed by methanol. Now mix it well to dissolve the mixture. The stain so prepared must be kept as it is and utilized the following day making sure that the moisture does not enter in the complete process.

Lugol Iodine solution -

Prepare from 1gm iodine, 2gm potassium iodide and 100ml distilled water. First we dissolve potassium iodide in 30ml of water and then add iodine to it. Mix the solution till the time it is completely dissolved and then add 70ml water to it. After mixing it well the solution is to be stored in a dark brown bottle.

Methylene Blue Aqueous -

This is a solution for Zehl Neelsen stain and obtained from mixing 0.3gm of methylene blue in 100ml of water. The solution is filtered and stored in a bottle.

Orthotolidine Reagent -

There are two solutions used in making the reagent- A&B.

Solution A - obtained from mixing 1.35gm of orthotolidine dihydrochloride in 500ml of distilled water.

Solution B - prepared by mixing 150ml of concentrated HCl with 350ml distilled water.

Now both the solutions A & B are mixed with continuous stirring and the solution so obtained is stored in amber colored bottles.

RBC Diluting fluid -

It is prepared by mixing 3gm sodium citrate, 1ml commercial formaldehyde solution containing minimum 37% formalin and 100ml distilled water.

Safranine solution -

In order to prepare the working safranine solution, first the stock solution is prepared by mixing 2.5gm safranine O and qs 100ml ethanol 95%. Once this stock solution is ready then the working solution is made by adding 90ml distilled water to 10ml of the stock solution.

Sodium Chloride solution (0.85%) -

This solution is also referred to as isotonic saline or normal saline and obtained by dissolving 0.85gm sodium chloride in 1000ml distilled water.

Sodium Thiosulphate Aqueous solution -

This is used for neutralizing any chlorine in water samples which are taken for bacteriological examination. It is prepared by mixing 3gm sodium thiosulfate with qs 100ml distilled water.

WBC Diluting fluid -

The ingredients required are 4ml glacial acetic acid, 10 drops of aq methylene blue solution and qs 200ml distilled water. Firstly mix the glacial acetic acid with water and then add 10 drops of aq methylene blue to it.

Wills Solution -

It is also known as the saturated solution of sodium chloride. This is obtained by mixing 125gm of NaCl with 500ml distilled water by heating it to the boiling point. The mixture is then left to cool down. Once it is cold then see that there is some salt powder left undissolved and in case it is not then add 50gm more NaCl. After that filter the solution and store in a corked bottle.

Exercises:

What are types of analytical balance?

How will you weigh an object in chemical balance?

Give a detailed note on maintenance of balances.

How do you dilute reagents?

Explain in detail the guidelines of reagent preparation.

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