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Thalassemia is best thought of as a group of disorders rather than a single disease. They demonstrate a hemoglobin synthesis disorder in which there exists a defect in the rate of production of one or more of the globin chains. This defect results from either a heterozygous or homozygous deletion or inactivation of a globin chain gene.

Gene deletions that cause alpha thalassemia can be homozygous or heterozygous deletions. Homozygous alpha thalassemia (alpha thalassemia major), also known as hydrops fetalis, is a lethal hemoglobin disorder which usually results in stillborn infants. Both alpha chain loci on each chromosome of the pair are deleted, resulting in a total absence of alpha chains. These chains are needed for all normal hemoglobins. If born live, infants with alpha thalassemia major exhibit hepatosplenomegaly, ascites, edema, low birth weight and die within a few hours. Ethnic groups most commonly associated with this form of alpha thalassemia include primarily Southeast Asians and sometimes people of the islands in the Mediterranean.

In the homozygous state (-/-), both parents contribute one missing locus.(drawing modified from Harmening, 1999)

Look at the phase in which reactions are occurring. Reactions at immediate spin (IS) usually are not clinically significant. Reactions at AHG are clinically significant. Check for a match in the reactivity pattern by comparing sample reactions and individual antibody reactions Varying strength of reactions could indicate dosage. Dosage means that there are two "doses" of the same antigen present on the red cells . Antibodies that exhibit dosage react more strongly with homozygous cells (e.g., Jka Jka ) than with heterozygous cells (e.g., Jka Jkb) .

Cells 4 and 9 may be used for rule outs due to negative sample reaction. Screen cell I may be used for rule outs due to negative sample reaction. Look at the antigens present on cells 4 and 9 that are in the homozygous state (highlighted in green). Remember the 3 to rule in and 3 to rule out procedure. Antibodies ruled out (with 3 reactions): e, k, Kpb, Jsb, Jka, Leb, P1, Lub. A selected panel should be set up to rule out (with 3 reactions) the remaining clinically significant antibodies (E, D, C,c, K, Fya, Fyb, Lea, M,N, S, and s).

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

The selected cells should be antigen-negative for the antibody that you think is present and antigen-positive (homozygous) for what you are trying to rule out. You are designing a panel that addresses your testing needs. Example: JkbIf you suspect that your patient has an anti-Jkb and further rule out cells are needed, then those rule out cells should be negative for Jkb. In the table below, donor cells 1,2, 4, 6, 9 and 10 may be used when creating a select panel to test the patient and help rule out the remaining possible antibodies. The homozygous rule applies when choosing which cells to use for testing (antigens highlighted in light-yellow).Example: Picking cells to rule out CUse panel cell 1 and panel cell 2 (C is in the homozygous state). Explanation: Panel cells 1 and 2 do not contain the antigen Jkb (signified by "0" on cell panel). If these cells are tested with the patient's plasma and the reaction is negative, it can be assumed that the patient does not have an antibody to C. C is now ruled out because there would be a total of 3 negative patient reactions with C positive cells (These two reactions and screen cell I from the antibody screen, shown again below). This should be done for all clinically significant antibodies that you were unable to rule out on the first panel.

Start with the first panel cell where you have negatives in all the phases tested. Use the cells where patient reactions are negative in all phases tested for rule outs. Look at the screen cell antigram to see which cells the patient reacted with. You may use a screen cell as a rule out cell if there were no reactions in any of the phases tested and if it is homozygous for the antibodies you are trying to rule out; antigens should be in their homozygous state in order to rule out. Refer to the screen cell antigram example below to see homozygous screen cell that can be used for rule outs. Write down what you could not completely rule out with 3 homozygous cell reactions. If Jkb is the suspected antibody, then reactions with screen cell I should be negative. This screen cell may be used as a rule out cell.

Purpose: To design a set of panel cells that may help you to rule out additional antibodies and lead to the identification of the antibody that is present in the patient's plasma.Benefit of running selected cell panel: Decreases the use of reagents and specimen. How to choose selected panel cells: If you suspect that a specific antibody is present, the cells you choose for the select panel should be negative for that antigen and positive for the antigen you are trying to rule out (homozygous state).

Rule-out (also referred to as exclusion or cross-out) is a process by which antibodies are identified as being unlikely in a given sample because of the absence of an expected antigen-antibody reaction. In other words, the absence of a reaction is noted with a cell that is positive for the corresponding antigen. Non-reactive cells are selected for rule-out. To be classified as non-reactive, a cell must NOT have reacted in any phase of testing in a given panel or screen. In the case of cold antibodies: if reactions are only occurring at immediate spin and are negative in the AHG phase, then that panel cell can be used as a rule out cell for IgG reactive antibodies but not for antibodies that react at immediate spin (IgM).If there is no reaction with a panel cell then it is possible that antibodies to the antigens on that cell are not present in the sample being tested. Based on Fisher's statistical probability recommendation, the probability of having reliable results increases if you are able to have more rule-out and rule-in cells. By comparing the patterns of reactivity and non-reactivity, we can more safely assume that an observed pattern is not the result of chance alone. If a "3 (reactions) to rule in and 3 (reactions) to rule out" protocol is used, there is then a 95% probability that the reaction pattern is not due to chance alone. Homozygous cells are used so that weaker reacting antibodies which fail to react to the antigen present in the heterozygous state aren't accidentally ruled out. Examples of Homozygous and Heterozygous Antibodies Jka Jkb Patient IS Patient AHG Panel cell 10 + + 0 2+ Panel cell 11 0 + 0 4+ Panel cell 10 shows Jkb in the heterozygous state. The patient's reaction is weaker than the reaction with panel cell 11 which shows Jkb in the homozygous state.Reactions are weaker when antigens are present in the heterozygous state because there is less of the antigen present for the potential antibody to bind with.

Thalassemias are part of a group of hemoglobin synthesis disorders in which a defect exists in the rate of production of one or more of the globin chains. This defect results from either a heterozygous or homozygous deletion or inactivation of a globin chain gene.Thalassemias are named according to the affected gene or the globin chain that is showing reduced or absent synthesis.Globin chain loci are found on: chromosome 11 (beta, delta, epsilon, and gamma) chromosome 16 (alpha, and zeta)

Delta-beta thalassemia exists in both heterozygous and homozygous forms. The symptoms are mild to moderate depending on the severity of the disease.This form of beta thalassemia can be found in many ethnic groups, but is most common in persons from Greece and Italy.

The prevalence of common HFE mutations among persons with hereditary hemochromatosis (HH) has been reported in numerous studies conducted in the US, France, Australia, and other countries. Homozygous C282Y mutation (C282Y/C282Y) is present in 82% to 90% of Caucasian patients diagnosed with iron overload due to HH.(7) This suggests a strong link between the genotype and the phenotypic presentation of clinical iron overload. Much lower percentages of persons diagnosed with HH do not have two C282Y mutations. A small percentage of persons diagnosed with HH are compound heterozygotes for C282Y and H63D (C282Y/H63D), are homozygous for H63D (H63D/H63D), heterozygous for C282Y (C282Y/wild type) or for H63D (H63D/wild type), or carry S65C or other HFE mutations.It may be that symptomatic heterozygotes are actually HFE-compound heterozygotes with additional unidentified mutations modifying the expression of the more severe known mutation. It is quite possible that more mutations of HFE and elucidation of other gene mutations modifying HFE will be discovered in the future enabling scientists to better explain the phenotypic heterogeneity of this disorder.In the US, the C282Y mutation is most prevalent in the non-Hispanic white population. It is much less common among Hispanics and African Americans.

For reasons as yet unknown, not all individuals who are homozygous for the C282Y mutation display phenotypic features of HH, and persons with H63D polymorphisms rarely develop iron overload. The penetrance (percentage of individuals with a specific genotype who express the associated phenotype) of HFE mutations is generally considered to be low. Results of a recent meta analysis by the US Preventive Services Task Force conclude that 38% to 50% of all C282Y homozygotes develop some evidence of iron overload, but that only 10% to 33% develop clinical disease due to HH. (8) In other words, some individuals may have elevated iron test results such as transferrin saturation, but do not demonstrate significant organ damage. Estimates of penetrance in some studies have found it to be even lower. Penetrance of HFE mutations is currently a controversial subject among experts, and the significance of finding HFE mutations in a given individual is often unclear. The probability that a given individual with HFE mutations will develop clinical disease from iron overload cannot be determined at this time.

Homozygous “hh” individuals do not form H substance and thus have no way for late sugars to attach. The blood group resulting from the homozygous “hh” condition is called the Bombay blood group (Bombay phenotype). Due to the presence of anti-H in the serum of a person with the Bombay phenotype, only blood from another person with the Bombay phenotype may be transfused.

The family (cited in the previous case history) was from a region of Thailand where the physician knew HbE carriers are prevalent. Homozygous hemoglobin E is common in Southeast Asia and presents with very mild anemia and seldom requires transfusion. Over 30 million people in the world are HbE carriers, making this abnormal hemoglobin almost as common as HbS. Hemoglobin E is uncommon in North America and in Europe, but with changing immigration patterns, hemoglobinopathy E cannot be ignored. Peripheral blood smear findings of target cells, microspherocytes, red cell hypochromia, a few red blood cell fragments, and nucleated red blood cells require evidence from hemoglobin electrophoresis to establish a diagnosis. Clinically, a very important and severe syndrome is hemoglobin E/beta thalassemia in which there is hemolysis requiring repeated transfusions. The patient has a severe anemia, low MCV (50's), and high RBC. This is characteristic of Hgb E/beta thalassemia.

Sickle cells can be seen in the peripheral blood of patients who have homozygous sickle cell anemia; however other tests are needed to make the diagnosis. Most sickled cells can revert back to the discoid shape when oxygenated. About 10% of sickled cells are unable to revert back to their original shape after repeated sickling episodes.

Pelger-Huet anomaly is the inherited form of neutrophilic hyposegmentation. Its transmittance is autosomal dominant and the anomaly is present in one out of 6000 people. When present, all of the neutrophils will be hyposegmented; however, the homozygous state will have increased number of cells with singular round nucleus and decreased numbers of the bilobed forms.

The Pelger-Huet anomaly is a congenitally acquired condition of nuclear segmentation and is clinically insignificant. There is no loss of cellular function.The condition can be suspected if typical bilobed, "pince-nez" nuclei are observed (left upper frame in the composite photograph).Band neutrophils usually have two distinct lobes, connected by a relatively short but thick bridge as illustrated in the upper and lower right frames. Monolobated cells may also be encountered, as illustrated in the lower left frame. If these are seen in significant numbers, the possibility of a homozygous Pelger-Huet should be considered.