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A 42 year old male received 6 units of RBCs during an open heart surgery 6 months ago. His antibody screen was negative at that time. He has returned for a follow up surgery and his antibody screen is now positive with both screen cells at the AHG phase.Reactions are occurring at AHG phase which indicates a possible clinically significant antibody, Jka showing dosage. Refer to Case Study 1 panel below to see reactions of antibody panel.IS = Immediate Spin; AHG = Antihuman Globulin Phase; CC = Check Cells; AC = Auto Control; ND= Not doneCase study 1 conclusion:Patient's previous transfusion 6 months ago exposed him to the Jka antigen, causing the formation of this antibody which is known for showing dosage.

Possible antibody is anti-C based on matching reaction pattern of sample at AHG. At least 3 positive reactions are present to rule in this antibody.Pink: negative reactions to use for rule-outsTurquoise: homozygous reactions used for rule-out (exceptions to homozygous rule are Rh group and Kk) Antibodies that can be ruled-out using "3 to rule out" rule: D, c, E, e, K, k, Fya, Fyb, Jka, Jkb, Lea, Leb, M, N, S, s, P, LubAntibodies that cannot be ruled out: Cw, Kpa, Jsa, LuaPoints to remember: The pattern of positives and negatives on an antibody panel cell indicates whether that particular antigen is present on the testing cells The phase in which the reactions are occurring will help determine if it is an IgG clinically significant antibody or IgM antibody (usually not considered clinically significant). Stronger reactions seen if antibody exhibiting dosage. Think multiple antibodies if reactions occurring at different reaction phases.

In this example the patient's plasma tests positive with both screening cells at a strength of 4+. In the panel below, reaction patterns show varying strengths, 2+ to 4+ (highlighted in green).4+ could indicate one strong antibody or a combination of several antibodies that increases the strength of the reaction.3+ could indicate one strong antibody.2+ could indicate one the reaction between one weak antibody and the corresponding antigen that is present on with the other target antigen not present on that testing cell. If the panel cell is in the heterozygous state, the reaction of the antibodies present may be weaker if they commonly exhibit dosage. Since Cw, Kpa, Jsa, Lua are not present on the testing cells they are probably not causing these reactions. Perform rule outs using panel cells 5 and 7 (sample had no reaction in any phase with these panel cells) Cells that have at least 1 out of the 3 rule outs needed: C, c, e, K,k, Kpb, Jsb, Fya, Jkb, Lea, M, N, s, P1, Lub Antibodies that could not be ruled out with this panel: D,E, K, Fyb, Jka, Leb, S Predominant pattern of 4+ in panel cells 1,2,3,4,10 matches D Varying strengths in reactions indicates a possible second antibody so selected cells should be picked to aid in identification Find a panel cell negative for D (antibody you suspect) and homozygous positive for the antibody you are trying to rule out. For example: D E e K k Fya Fyb Jka Jkb Lea Leb S s Donor cell 1 0 0 + 0 + 0 + + + 0 + 0 + Donor cell 1 could be used as a rule out test for e, k, Fyb and Leb. Reactions should be negative if these antibodies are not present.You should have a total of 3 negative reactions with panel or screen cells to rule out potential antibodies. If reactions with this panel cell are negative, then e and k can be ruled out with a total of 3 to rule out reactions. Selected cells should be picked for each antibody that needs to be ruled out in order to determine the identity of the other antibody

Suspect dosage if varying strengths in reactivity are seen and reactions are in the same phase. Weaker reactions will be seen if suspected antibody is reacting with antigens in the heterozygous state. Stronger reactions are seen if the antigen is present on the testing cells in the homozygous state. This allows more corresponding antibody to bind with the antigen. Remember the antibodies known for showing dosage are: Rh, Kidd, Duffy, MNSs, and Lutheran. Dosage may be seen if cells are R2R2 (DcE/DcE). These red cells have more D antigen sites so reaction with anti-D may be stronger.Refer to Example 5 on the following page.

Ketone bodies are usually absent in urine. The presence of ketones in the urine probably indicates that the body is using fats rather than carbohydrates for energy. High levels of ketones may be present in the urine of individuals with uncontrolled diabetes because the body's ability to metabolize carbohydrates is defective. Detecting the presence of ketones in the urine is a valuable aid to managing and monitoring individuals with diabetes mellitus. Ketonuria is an indication that the insulin dose needs to be increased. Electrolyte imbalance and dehydration occur when ketones accumulate in the blood. If these conditions are not corrected by adjusting the dose of insulin, the patient may develop ketoacidosis and ultimately diabetic coma. Low levels of ketones may be detected during conditions of physiological stress such as fasting, rapid weight loss, frequent strenuous exercise or prolonged vomiting. The presence of ketones in these situations is due to either inadequate intake of carbohydrates or increased loss of carbohydrates.

The use of heparin is prophylactic. It is used either to prevent thromboembolism (a condition in which a blood clot forms inside a vessel), or used to limit a previous thromboembolism. Heparin inhibits thrombin. The degree of inhibition is dosage dependant. Low doses of heparin inhibit initial thrombin formation in the coagulation cascade, and act to slow down overall thrombin generation. At higher doses, heparin can inhibit thrombin entirely, making blood coagulation impossible. Heparin is a potent anticoagulant. Accurate monitoring is essential. The activated partial thromboplastin time (APTT) and/or activated clotting time is used to monitor unfractionated heparin therapy.

Adverse drug reactions are a leading cause of morbidity and mortality in the United States. Drug metabolism is a process whereby drugs are delivered to the body, distributed, metabolized and then ultimately excreted. As there can be potentially significant differences between patient drug absorption, metabolism and excretion, molecular testing allows a physician to work with a patient's individual phenotype and/or genotype to deliver an optimum pharmaceutical selection and/or dosage.

TDM provides a quantitative measure of the circulating concentration of a drug. The physician determines if the dosage of the drug needs to be adjusted based on this information.If a drug concentration is determined to be outside the therapeutic range, it may be for one of the reasons listed in the table below. Reason Discussion Noncompliance Patients may (intentionally or unintentionally) not take the drug. TDM can thus help monitor compliance. Dosing errors The dose may have been erroneous or inappropriate given the patient's condition. Malabsorption The TDM result will reveal if the drug cannot be absorbed well through the gut and an alternative route of administration will be needed. Drug interactions Many drugs interfere with the absorption or metabolism of other drugs. These interactions will be revealed by TDM. Kidney or liver disease Any pathology that affects elimination will cause an elevation in a drug level that will be unmasked by TDM. Altered protein binding Changes in serum proteins can lead to big changes in the amount of free drug in serum. Variations in the genetics of drug-metabolizing enzymes can also affect drug concentrations in the body. This is the field of pharmacogenomics that will be discussed later in the course.

Similar to evaluating inconsistencies, one of the post-analytic tools for confirming that the serological data fit the solution is to consider the "big picture." For example: Is there a likely red cell stimulus (prior transfusion or pregnancy) for IgG antibodies such as anti-Jka? Can different reaction strengths with panel cells be explained by the identified antibody (e.g., dosage) or by the presence of more than one antibody? Is the antibody unusual for a patient of a particular race? For example, anti-Dib is more likely to occur in Native Americans than in Caucasians.

The antibody in the pretransfusion specimen (prior to the patient being transfused with two units of unmatched group O Rh-negative RBC) reacted 2+ and 3+ with antibody screen and donor cells.If Jk(a+), the transfused donor RBC would have stimulated increased antibody production and the patient's plasma would be expected to react even more strongly with Jk(a+) red cells than in the pretransfusion specimen.However, the expected increase in antibody strength did not happen. Because Jk(a+) donor cells "mop up" (adsorb) the patient's anti-Jka, initially the anti-Jka decreased in strength. Later, once donor red blood cells are no longer present to adsorb the antibody, the anti-Jka would be expected to become stronger.Currently, (2-weeks post-transfusion) the patient's plasma is only reacting 1+ with Jk(a+b-) RBC and w+ with Jk(a+b+) RBC.This effect is called dosage. Learning points When a secondary immune response occurs, antibody first decreases before it increases. The expected increase in antibody strength will vary depending on the amount of excess antibody available in the patient's plasma at the time of testing versus the amount that had adsorbed to donor rbc and been removed by EVH.~