• Join over 1.2 million students every month
  • Accelerate your learning by 29%
  • Unlimited access from just £6.99 per month

Immunology Practical: Enzyme Linked Immunosorbent Assay (ELISA)

Extracts from this document...


Immunology Practical: Enzyme Linked Immunosorbent Assay (ELISA) Aim To acquire understanding and knowledge to determine the antibody titre of the Rabbit Anti-Ferritin Antibody using the Enzyme Linked Immunosorbent Assay (ELISA) Introduction ELISA is a rapid immunochemical assay that involves an enzyme (a protein that catalyzes a biochemical reaction). It also involves an antibody or antigen (immunologic molecules). The ELISA is a fundamental tool of clinical immunology, and is used as an initial screen for pathogens detection. Based on the principle of antibody-antibody interaction, in this assay you can easy visualisation the results and can be completed without the use of radioactive materials. The ELISA technique is the first and most basic assay to determine if an individual is positive for a selected pathogen, such as HIV. The applications of immunoassays are extended to other fields such as infectious diseases, autoimmune diseases, cancer, degenerative diseases, haematology and pharmacology. Applications have not been confined to human health care. Many applications have been described in veterinary medicine, agriculture, environmental health and the food industry. The assay is performed in a microtitre plate (plastic) which contains an 8 x 12 matrix of 96 wells. They also called indirect ELISA or sandwich ELISA. Microtitre plates are coated with antigen. ...read more.


Control 100�l of diluting buffer 100�l of 2� Ab H Once you have added all the solutions into the wells, you then place the plate into the incubator at 37�C for 30 minutes again covering the plate. After the 30 minutes are over, you now wash the plate again with diluting buffer 3 times, but this time you are also using distilled water using the same methodology as before. Now you need to add 100�l of urea; the substrate to all the appropriate wells as mentioned in the table below: Table3 Well's Columns Well's Row 1 2 3 4 5 6 7 8 9 10 11 12 A Add 100�l of UREA (Substrate) 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " B 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " C D 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " E 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " 100�l " F G Control 100�l " 100�l " H You then incubate the plate for 24 hour at 37�C. ...read more.


In wells D3, 6, 7, 8 there is slight contamination. D3 D6 D7 D8 The reason why we know that this is a contamination is because there is no positive reaction in the first wells (D1 or E1; the highly concentrated wells). Limitations to ELISA: The drawback to the antigen-coated plate method is that it is precise; limiting quantity of antigen must be bound to the microtitre plate. Furthermore any dissociated antigen will also act as an inhibitor. Reference: 1) Definition of Enzyme-linked immunosorbent assay (ELISA) http://www.medterms.com/script/main/art.asp?articlekey=9100 [accessed on the 20 Nov 2008 at 13:52] 2) G.R. Bullock, D. Van Velzen, M.J. Warhol. (1991), Volume 2. Techniques in diagnostic pathology / edited by G.R. Bullock, D. van Velzen and M.J. Warhol. Vol.2, ELISA techniques: new developments and practical applications in a braod field / volume editor, P. Herbrink. London: Academic Press. 3) Introduction to ELISA Activity. The Biology Project, The University of Arizona, Wednesday, May 3, 2000. http://www.biology.arizona.edu/IMMUNOLOGY/activities/elisa/main.html. [Accessed on 27th November 2008]. 4) Kemeny, D. M. (1991). A practical guide to ELISA / D.M. Kemeny. Oxford: Pergamon 5) Lequin RM. Enzyme Immunoassay (EIA)/Enzyme-Linked Immunosorbent Assay (ELISA). Clin Chem 2005;51:2415-2418 6) The Wellcome Foundation Ltd, United Medical and Dental School of Guy's and St Thomas's Hospitals. (1988). ELISA and other solid phase immunoassays: theoretical and practical aspects / edited by D.M. Kemeny and S.J. Challacombe. ChichesterbWiley. ?? ?? ?? ?? - 1 - ...read more.

The above preview is unformatted text

This student written piece of work is one of many that can be found in our University Degree Cell Biology section.

Found what you're looking for?

  • Start learning 29% faster today
  • 150,000+ documents available
  • Just £6.99 a month

Not the one? Search for your essay title...
  • Join over 1.2 million students every month
  • Accelerate your learning by 29%
  • Unlimited access from just £6.99 per month

See related essaysSee related essays

Related University Degree Cell Biology essays

  1. The purpose of this investigation is to discover whether different respiratory substrates will affect ...

    Controlled Variables Controlled Variable How I will control it Why I will control it Temperature By placing the boiling tubes in a water bath that is heated constantly at 40C The temperature must be controlled because the temperature will affect the rate of respiration of the yeast.

  2. This experiment was carried out to characterize an enzyme, -amylase by extracting it from ...

    was also determined after a series of purification to purify the enzyme. Results: a) Protein determination of ?-amylase Table 1: The absorbance and mass of protein in standard BSA and ?-amylase crude extract at wavelength of 560nm Solutions Absorbance 1, 560nm Absorbance 2, 560nm Mean absorbance, 560nm Mass of protein,

  1. Proteins. The experiment aimed to extract proteins from green papaya through salting out with ...

    The fractions that were collected contained the maximum protein concentration and minimum salt concentration. On the other hand, figure 3 shows the SDS-PAGE gel used in the experiment. The first and third well from the right contained the crude

  2. Skin Cancer

    Additionally, each form of skin cancer has distinguishing characteristics. Basal cell carcinoma could appear as a sore that grows slowly, yet does not itch or hurt. It may have a dimpled middle or rounded edges. Typically, basal cell carcinoma will appear in an area with sun damage. There is no conventional size or shape for it to be.

  1. IMMUNOLOGY PRACTICAL: Differential Blood Cell Counts

    The stains used will stain your skin and are also potentially harmful if ingested. Standard laboratory procedure applies. Staining of Blood Smears; fix the cells on the smear in 95% methanol for 2 min. then Immerse the slide in buffer for 5 min (remember which side of the slide the cells are on).

  2. Biology Light & Life

    - Photon of light is absorbed >> the retinal pigment molecule changes shape >> alterations to the opsin protein are triggered >> more downstream events are triggered: alteration in intracellular ion concentrations and electrical signals. The electrical signals are sent to the visual centres of the brain.


    To me it seems that any strict definition of life which differentiates the living from the non-living represents a too binary model which is not supported by empirical evidence available. However, whilst it is very difficult to define life, a particularly interesting characteristic of life as we know it is

  2. Discuss how changes in control of the cell cycle contribute to cancer development ...

    Checkpoints are intricate sensor mechanisms within the cell which monitor the cellular environment and determine whether appropriate conditions have been fulfilled before it progresses further through the cell division cycle (Beck, et al., 2009). Ergo a major function of these checkpoints is to oversee that the integrity of the genome remains intact throughout the cell cycle.

  • Over 160,000 pieces
    of student written work
  • Annotated by
    experienced teachers
  • Ideas and feedback to
    improve your own work