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A biotechnology scientist must execute his/her Laboratory techniques successfully. The success of the experiment depends merely on the truth in executing the experiment. And Accuracy can be achieved by the experience and besides depends on cognizing the proper cognition about the instrument use and detecting the proper techniques involved in the executing of the experiment and the instruments. And some of the common research lab techniques are discussed below, in the techniques we must detect the usage of analytical balance, the use of the pH metre and cognizing how to graduate, microcentrifuge working, micropipette use and really of import labelling the reagents decently.

Preparation OF A BUFFER REAGENT

In this experiment with the aid of the analytical balance, micropipette and pH metre buffer reagent must be accurately prepared. The buffer reagent consists of 0.1M Na chloride and 0.025M citric acid with pH of the buffer solution is maintained at 6.0. The weights of the chemicals must be weighed by utilizing the analytical balance carefully as the instrument is really sensitive. And pH of the solution is maintained by utilizing the pH metre by plunging the pH metre electrode into the solution and observing down the reading displayed by the pH metre on a regular basis. The aims of the experiment is to fix 100ml of stock buffer solution in a container which consists of 0.1M of Na chloride and 0.25M of citric acid with the pH-6 and to cognize the proper use of the analytical balance and standardization of the solution pH accurately by utilizing pH metre.

Materials

100ml beaker is needed for blending the chemicals ; disposable pipettes are used for commanding pH seting bead by bead of 3M NaOH for levelling the pH. pH metre with pH electrode is used for mensurating the pH by puting it in buffer reagent prepared by utilizing 0.1M Sodium Chloride, 0.25M citric acid monohydrate which are measured utilizing Analytic balance. Extra containers required are volumetric flask 100ml and 100ml screwcap bottles they must be washed exhaustively by utilizing deionised H2O nowadays in the wash bottle. Standard buffer, Spatulas and baseball mitts are besides required.

Method

The sums of chemicals are required for the readying of 100ml solution can be found out by utilizing molar concentration equation and known as 0.025M citric acid and 0.1M NaCl.

_Mass of solute ( gm ) /FWsolute_

M = Volume of solution ( litre )

The chemicals are weighed as deliberate and transferred to the 100ml beaker and they are dissolved by utilizing 90ml of the distilled H2O. pH metre is calibrated by utilizing standard buffer solutions on is of pH 4.0 and the other is pH 10.0 at the room temperature.

The solution in the 100ml beaker ( NaCl + Citric acid ) is assorted good and the graduated pH metre is placed inside the solution and the mixture pH is easy and carefully adjusted to 6.0 by adding 3M NaOH bead by bead with the aid of the pipette and continuously stirring the solution.

After the solution has reached to the needed pH that is 6.0 the solution is transferred to the volumetric flask of 100ml the volume is measured precisely by seeing the measuring Markss provided on the surface of the flask.

And the solution is transferred to the screwcapped nalgene bottle to hive away the solution for the hereafter usage and bottle with the solution is decently labelled for clear designation and indicant.

Consequence

_Mass of solute ( gm ) /FWsolute_

M = Volume of solution ( litre )

.

Gram = M – FW – volume of solution

Sum of NaCl= NaCl ( FW- 58.44 )

Gram = 0.1-58.44-0.1

= 0.5844

Sum of Citric Acid = citric acid ( FW- 210.14 )

Gram = 0.025 – 210.14 – 0.1

= 0.5253

Carefully 3M NaOH is added bead by bead at changeless stirring to set the pH of the solution to 6.0.in the 100ml beaker.

First before computation the units must be converted 1mM = 0.001M and 1ml = 0.001litre

Gram = M – FW – volume of solution

NaCl = 58.44 – 0.25 – 0.14

=2.0454

KCl = 74.55 – 0.25 – 0.004

= 0.07455

NaH2PO4.H2O = 137.99 – 0.001 – 0.25

=0.03449

MgSO4 = 120.37 – 0.0009 – 0.25

=0.0270

HEPES = 283.31 – 0.01 – 0.25

= 0.7082

Glucose = 180.2 – 0.025 – 0.25

= 1.1262

All the reagents are taken in to the 250ml of flask and fade out utilizing distilled H2O, now set the pH to 7.4 utilizing 2M NaOH in the flask utilizing dropper or pipette.

Discussion

Accurate mixture of the chemical composings will give the accurate consequences so we must be careful while weighing the chemicals and blending them n the proper proportions. One must take attention of while he/her covering with the NaOH as it is really caustic we must follow some of the wellness and safety safeguards like have oning baseball mitts and goggles when they are managing the chemical.

The pH metre consist of a pH sensitive bulb which is made up of a really thin glass and it is really delicate 1 must be really careful while managing the equipment. The glass bulb may check even if it touches the bottom surface of the beaker while proving the pH of the solution. Before utilizing the pH metre it must be calibrated, and we must ne’er let the pH electrode to acquire dried up. It must be stored by plunging it in storage solution like 4M KCl solution with pH 4 or 7 buffer solution. While we about to utilize it, it must be taken out of the storage solution wipe it and graduate it with the standard solution, wipe the bulb once more and plunge it in to the trial solution look into the reading and once more hive away it in the storage solution.

Analytic balance is a really sensitive device and it is digital it has a capableness of weighing the substance up to 0.0001 or even 0.00001 gms and it is really accurate. It must be placed in the prohibitionist and the clean country and the country must by degree. Over lading the balance more than its capacity may botch the device. The excess weight of a home base or a paper placed on the balance can be tared. And the safeguard must be taken that the air besides must non come in in the device while mensurating, doors of the device must be closed to derive truth.

Labeling is really of import on the storage bottle as it tell the whole information about the solution that contains in the bottle the label must hold the information about the chemical composing, pH of the solution, when it is made and the batch figure if it is made by a group. So that it will be easy for the designation for the future us of the solution for any other experiment.

Mentions

Barry Haggett, 2010. Introductory experiment on common research lab instruments. Learning resources, Analytical and Diagnostic Technologies. [ Online ] . Available at: hypertext transfer protocol: //breo.beds.ac.uk ( Accessed: 26 May 2010 ) .

CETRIFUGATION OF SUSPENSIONS

Centrifugation is most widely used technique in different field of research like Biochemistry, Cell and Molecular biological science and in the medical specialty. The technique is used to insulate the cell organelles inside the cell. Centrifugation procedure is nil but whirling the samples at really high velocity, which result in the separation of the atoms based on their denseness due to the gravitative force and the precipitation will be formed in the underside of the tubing which is besides called as pellet. And the staying solution on the top of the pellet is called as the supernatant. After separation they must be instantly separated by utilizing the micropipette without upseting the pellet. The extractors are two types among which are used in the research lab intent. One is the simple extractor which is regular size incorporating regular size extractor tubings or centrifuge tips and the other one is micro extractor which consists of micro extractor tips. They both different in their velocity and capacity, the rate of the centrifugation depend on the acceleration that is applied to the sample.

The rotor consists of a covering palpebra which is interlocked to originate the spinning of the rotor and acts as a defender from any touch injures during the procedure. And besides from the inadvertent catastrophically rotor fails. The rotor must be ever balanced, samples inside the rotor are placed such a manner that they lie opposite to each other with equal multitudes. If the empty infinite is available on the antonym of any sample it must be filled with H2O sample of same mass to equalize the balance of the rotor.

The acceleration rate of the extractor is measured in footings of revolutions per minute ( RPM ) or RCF ( comparative centrifugal force ) and unit of gravitation as ( g ) . Some processs of separation atoms and liquid are based on the centrifugal status which is specified in footings of RCF or unit of gravitation but most of the micro centrifuges consist of puting for merely RPM. So the relationship between RPM and RCF is

g = ( 1.118 x 10a?»a?µ ) RS?

Where g = RCF, R is the radius of the rotor in centimeter and S is the velocity i.e. RPM.

The aims of the experiment are to cognize the usage of the micropipette to mensurate the volume and pouring liquid, use of Micro extractor and change overing RPM to RCF.

Materials

1000µl and 200µl micropipettes are needed with at least 60 micropipette tips for measuring, 1.5 milliliter micro extractor tubing for puting the samples in it to run micro centrifuge 70 in figure at least. 2 micro extractors for the procedure, try A and B incorporating black and green coloured mixtures severally and a swayer to mensurate the radius of the rotor.

Method

By utilizing 1000µl micropipette take 1000µl of the will blend sample for sample A and transportation into the 3 separate 1.5ml micro extractor tubings and put them in the rotor in the balanced status.

Measure the radius of the rotor with the aid of the swayer placing at the centre of the rotor to the border of the rotor and note it down the reading and cover the palpebra of the extractor. Put the extractor machine for 1 min at the velocity of 5000RPM and press the start button. After the extractor machine halt after 1min remove and fling the supernatant liquid carefully without upseting the pellet with the aid of the 200µl micropipette.

Now transfer 1500µl of sample from sample B into 2 separate micro centrifuge tubing in the same manner as explained antecedently and put them inside the rotor in the balanced status like opposite to each other. And set the extractor machine for 1 min and at the velocity of 5000 RPM. And look into if there is any separation or non, if the separation is non seen so increase the velocity to 10000RPM and take and fling the supernatant liquid carefully.

Calculate the RCF in footings of g from rotary motion velocities of the both samples.

Consequence

For the suspension A ( Blue coloring material )

g = ( 1.118 – 10-5 ) RS2

= 1.118 – 5 – 5000 – 5000 / 10000

= 1397.5

For the suspension B ( off Green coloring material )

g = ( 1.118 – 10-5 ) RS2

= 1.118 – 5 -10000 – 10000 /100000

= 5590

Discussion

Micro extractor is most widely used in the research labs for the isolation of the cell cell organs and macro molecules from the cells it is really simple procedure.

Care should be taken when puting the micro extractor tubings in the rotor they must be arranged in the balanced conditions otherwise the machine will do coggling sound and there is a opportunity of firing inside the machine and it may acquire spoiled. If u heard coggling sound signifier the machine instantly push the stop button and topographic point the tubings inside balanced and turn it on once more.

If there is no precipitation formed inside the tubing alteration the velocity of the rotary motion RPM ( increase the RPM ) . This can be the ground for the uncomplete separation of the precipitation.

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