Downloaded 12 times
ABO_Blood_Typing[1]
- 1. A Simulated Experimentfor ABO Blood Typing Amanda J. Powell Department of Chemistry, Bowling Green State University, Bowling Green, OH 43403; *powelaj@bgsu.edu Dr. Stephania Jo Messersmith Department of Chemistry, Bowling Green State University, Bowling Green, OH 43403; *messers@bgsu.edu Abstract One of the greatest challenges of teaching the concept of blood typing is the incorporation of common experimental techniques into the teaching laboratory. Instructors will often run into problems with finding both legal and environmentally safe chemicals to teach popular practices used by forensic scientists, biology technicians and others who use this type of testing. This laboratory exercise will specifically focus on using practical methods to teach students about the concept of blood typing by eliminating the need to use real blood and utilizing reactions from common chemical demonstrations. This lab uses chemical reactions that have commonly been used as demonstrations in order to simulate blood typing—reducing the biological hazards. It also uses chemicals that can be disposed of down the drain after the solids have been removed. Background The concepts behind blood typing are essential to almost all science courses. Blood typing is used in almost every forensic and biology laboratory and it is critical that students understand the theories behind the procedures used in determining blood types. This experiment will focus on accomplishing this without using human blood. Blood typing relies on the interaction between cell-surface proteins on erythrocytes and antibodies. The antigens on red blood cells correlate directly with the blood type. Researching activities on blood typing gave detailed examples of the process behind agglutination and coagulation, these articles can be found in the supplied references1-5 . Antibodies, proteins specific for an antigen, are produced when the erythrocyte antigens are recognized as “foreign.” A response will be elicited that results in the clumping together of the red blood cells by the antibodies, a process that is referred to as agglutination. Specific antigens have corresponding antibodies. For example, antigen A will have proteins present on its surface that will recognize and adhere to proteins present in Antibody A. No clumping takes place if the antibodies do not recognize their corresponding antigen. Blood type O will have no antigen proteins present on its surface and therefore both Anti-A and Anti-B will be produced in response against it. Blood type AB is the opposite in that it has both antigens A and B. The immune system will not produce any antibodies against type AB. Table 1: Blood Types and Corresponding Antibodies and Antigens 1 BLOOD TYPE ANTIGEN ANTIBODY A A Anti-B B B Anti-A AB A and B Neither O None Anti-A and Anti-B
- 2. Experimental Overview Table 2: Chemicalsused in Simulated Blood Agglutination The purpose of this exercise is to demonstrate the concept of agglutination as relevant to determining a person’s blood type. We focus on using chemicals common to most laboratories in order to simulate the reaction between blood antigens and antibodies. This experiment allows instructors to teach the theory of blood typing without the dangers of actually using real blood. Chemicals In order to imitate the reaction between blood cell antigens and their corresponding antibodies, chemicals were chosen that gave similar results. Great care was used to ensure that the correct antibodies precipitated with the corresponding antigens and not with any others. The reaction for blood type A will be a chemical reaction for the production of Sterno, a gel, as found in Chemical Demonstrations: A Sourcebook for Teachers1 . Similarly, the chemical reaction for blood type B will be taken from the same source. We first focused on simulating the agglutination of blood type A with its corresponding antibody, anti-A. In order to imitate this reaction, a solution of calcium acetate was used to replace blood type A and 100% ethyl alcohol was used as anti-A. Similarly, we researched chemicals that would react to imitate the agglutination of blood type B with its corresponding antibody, anti-B. A mixture of Borax® detergent and aluminum sulfate was reacted with sodium bicarbonate to give the anticipated precipitate. High purity (98%) egg albumin was also tried in place of the Borax® and gave a similar result, a lower purity sample (62-88%) was tried but it did not work. The egg albumin is not recommended due to the high price compared to the Borax®. Simulation of the AB blood type was simply done by combining the mixtures used for the separate blood types A and B at a one-to-one ratio. The resulting mixture gave precipitate with the above-mentioned anti-A and anti-B chemicals. Water was used for blood type O, giving no reaction with either the anti-A or anti-B serums. Procedure The procedure for this experiment is straightforward. Similarly, the chemical reaction for blood type B will be taken from the same source. The “clumping” together of antigens due to antibodies will be simulated by simple chemical reactions that produce a solid-like precipitate. Food coloring is used for convenience and the sake of demonstration purposes. Red food coloring can be added to the blood serums and yellow to their corresponding antibodies. A problem that arose was the coloration of ethanol. It does not seem to pick up the food coloring well. The instructor will begin by first preparing the solutions of fake blood and serums before hand. First, start with blood 2 CHEMICAL VOLUME/MASS BLOOD TYPE A Calcium Acetate 17g Tap Water 100mL ANTI-B 100% Ethanol 2mL BLOOD TYPE B Borax® 1.0g Aluminum Sulfate 7.0g Tap Water 50mL ANTI-A Sodium Bicarbonate 1.5g Tap Water 150mL
- 3. type A. Asolution of saturated calcium acetate is prepared by dissolving calcium acetate in distilled water with stirring. This could take a few minutes. Anti-A is simply 100% ethyl alcohol. The exact masses and volumes of the mixtures can be found in the provided table. Blood type B is a solution of Borax® detergent, aluminum sulfate and water. After measuring out the given quantities of laundry detergent and aluminum sulfate, grind them both together in a mortar until the mixture appears as a fine powder with no obvious clumps. Dissolve this powder in warm water and then filter through quantitative grade filter paper. Dissolving sodium carbonate in water as well can make antibody B. To simulate blood type AB, a mixture of the solution used for type A and for type B was produced. Simply use a one-to-one ratio of the two solutions and use the antibodies as previously explained. Blood type O will simply be distilled water, eliciting no reaction from either antibody A or antibody B. After all of the solutions have been prepared, it is a good idea to label the beakers. For demonstration purposes, a drop of red food coloring was added to the blood serums and a drop of yellow food coloring was added to the antibody solutions. Students can begin the experiment either with blood type A or B. They will mix together the calcium acetate solution with 100% ethyl alcohol on a small watch glass or well plate. A clear gel like substance will precipitate out, imitating the agglutination of blood type A with anti-A. A similar reaction will occur with type B and anti-B but instead of a clear gel, white foam will be produced. Using a macro scale approach to this experiment will have the reactions looking like the clumping of erythrocytes. Blood type AB should react with both anti-A and anti-B, while type O will produce no reaction. Students should take care to record all results. Clean-Up As stated before, this experiment is aimed to be an environmentally friendly procedure, in which most of the materials can be disposed of by simply disposing down the drain, after the liquid is decanted from the solids. The solids should be collected for disposal and include calcium acetate and aluminum hydroxide.” Hazards Most of the chemicals used are common to most households, therefore there are a few risks associated with the materials. Ethanol is flammable and if inhaled, respiratory tract irritation can occur and it is known to cause skin irritation. For more information on any of the compounds used in this exercise, please refer to the appropriate MSDS information. Conclusion After vigorous research, it was found that very few, if any, experiments have been published that can successfully demonstrate the reaction between erythrocytes and their corresponding antibodies. The method outlined by Sharp and Smailes uses an experiment with a lead compound to imitate the agglutination of blood cells2 . Our experiment seeks to use chemicals with fewer health hazards and easier disposal methods. This experiment seeks to provide chemicals of low toxicity, and ones that are inexpensive as well. The reactions that occur do not fully resemble the actual reactions agglutination reactions of blood. However, this experiment is useful in simply demonstrating what takes place to entry- level students. It is a good hands-on experiment that better illustrates the 3
- 4. concepts associated withblood typing. This experiment can be used at the high school or college level. Note If watch glasses are not available, instructors can use small test tubes as well. This experiment is designed for a micro- level approach and thus, the quantities are small. To the authors’ knowledge, all current simulated bloody typing experiments use chemicals that are either toxic to humans and/or the environment3 . Literature Cited 1. Summerlin, L.R. (1985) Production of sterno: a gel. Chemical Demonstrations: A Sourcebook for Teachers, 134. 2. Sharp, J.D., & Smailes, D.L. (1989). How-to-do-it: a simulation of the blood type test. The American Biology Teacher, 51(4), 232-33. 3. Inheritance and blood typing. (2006). Vanderbilt Student Volunteers For Science, Retrieved from vaderbilt.edu/vsvs. 4. Summerlin, L.R. (1985) Production of a foam. Chemical Demonstrations: A Sourcebook for Teachers, 135. 5. Wake, C. (2005). Abo/rh blood- typing model: a problem-solving activity. The American Biology Teacher, 67(3), 158-62. 4