实验诊断学：LABORATORYEVALUATION OF ANEMIA

1、Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 1 1 1ST ST AFFILIATED HOSPITAL AFFILIATED HOSPITAL ZHENGZHOU UNIVERSITYZHENGZHOU UNIVERSITY Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 2 RBC disorde

2、rs (Anemias) Congenital disorders: Membrane, Hb & enzyme disorders. Acquired disorders: Decreased production Excessive destruction of erythrocytes Increased loss Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 3 Congenital RBC Disorders: (Hemolysis) 1. Membrane Disorders: Spherocyto

3、sis, Elliptocytosis 2. Hemoglobin Disorders: Hemoglobinopathies - Sickle cell, HbC etc. Thalassemia Syndromes - , , 3. Enzyme disorders: G6PD, PK deficiency 1 2 3 Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 4 Acquired RBC disorders : Decreased Production: Deficiency anemias Iron

4、, B12, Folate etc. Aplastic, Hypoplastic anemias Marrow disease, malignancy, radiation Increased loss/destruction: Blood loss anemias - parasites, bleeding Hemolytic anemias - Autoimmune (cold & warm antibody) mechanical, drugs & toxins. (Deficiency, Marrow disorder, Blood loss & Hemolytic) Liu, Yan

5、-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 5 LABORATORY EVALUATION Initial Hematocrit RBC Index Reticulocyte count (retic. count) Review peripheral blood smear Nucleated red cells, reticulocytosis Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 6 Pathological classific

6、ation of hemolytic anemia Extracorpuscular hemolytic disease (红细胞外在因素红细胞外在因素) Antibodies, infection, splenic sequestration and destruction, drugs, chemicals and physical agents, trauma to red blood cells. Intracorpuscular hemolytic desease (红细胞内在缺陷红细胞内在缺陷) Membrane, enzyme, hemoglobin Liu, Yan-Fang

7、MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 7 The site of hemolysis Intravascular hemalysis (血管内溶血) Extravascular hemolysis (血管外溶血血管外溶血) Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 8 LABORATORY EVALUATION Suspected Hemolytic Anemia Bilirubin-direct and indirect Lactate De

8、hydrogenase (LDH) Serum haptoglobin, urinalysis Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 9 LABORATORY EVALUATION Additional / Directed studies Coombs test Hams test, Sucrose hemolysis test, Rous Test Osmotic fragility test Hgb electrophoresis G-6PD assays Enzyme assays Bone m

9、arrow aspirate / biopsy Reversed M:E ratio Underlying disorders Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 10 Reticulocyte Count DEFINITION The reticulocyte is a young red blood cell which contains residual ribosomal RNA that can be stained with a supravital dye such as acridin

10、e orange or new methylene blue. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 11 Reticulocyte count Reticulocyte count gives an indication of the level of the bone marrow activity. Done by staining a peripheral blood smear with new methylene blue to help visualize remaining riboso

11、mes . The number of reticulocytes/ 1000 RBC is counted and reported as a %. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 12 Occurrence in blood* Adult: 0.5 - 1.5 % of erythrocytes New Born: 2 - 6 % of erythrocytes Absolute Count: (2484) 109/L * Measured by staining the blood smea

12、r. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 13 Anemia by Reticulocyte Index Impaired (Ret.) Hypoproliferation (AA) Appropriate (Ret. ) Hemolysis or Blood Loss 疗效观察 Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 14 Hematocrit Difinition The proportion of the

13、blood that consists of packed red blood cells. The hematocrit is expressed as a percentage by volume. The red cells are packed by centrifugation. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 15 Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 16 The Reference Rang

14、es Adult males: 42-49% Adult women: 37-48% Newborns: 55-68% One (1) week of age: 47-65% One (1) month of age: 37-49% Three (3) months of age: 30-36% One (1) year of age: 29-41% Ten (10) years of age: 36-40% Dependant on the age and, after adolescence, the sex of the individual. Liu, Yan-Fang MD, Ph.

15、 D. First Teaching Hospital ZhengZhou Univ. 17 What does a low hematocrit mean? A low hematocrit is referred to as being anemic. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 18 What does a high hematocrit mean? People living at high altitudes and in chronic smokers， Chronic cardi

16、opulmonary disease. Dehydration produces a falsely high hematocrit that disappears when proper fluid balance is restored. Some other infrequent causes of elevated hematocrit are lung disease, certain tumors, polycythemia vera, and abuse of the drug erythropoietin (18Epogen). Liu, Yan-Fang MD, Ph. D.

17、 First Teaching Hospital ZhengZhou Univ. 19 Mean Cell Volume (MCV) Definition Mean value Calculation The mean volume of each red blood cell. 80-100fl (1L=1015fl) hematocrit1015fl RBC (/L) MCV = Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 20 Mean Cell Hemoglobin (MCH) Definition

18、Mean value Calculation The mean value of Hb in each red blood cell. 26-32pg (1g=1012pg) hemoglobin1012pg RBC (/L) MCH = Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 21 Mean Corpuscular Hemoglobin Concentration (MCHC) Definition Mean value Calculation The mean concentration of Hb

19、in 1 liter RBC . 310350g/L Hemoglobin(g/L) Hematocrit (L/L) MCHC = Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 22 Anemia by Red Cell Indices Microcytic (95 fL) Megaloblastosis Liver Disease Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 23 Summary of MCV, MCH &

20、 MCHC SIGNIFICANCES AnemiaMCV MCH MCHCDiseases NormocyticN N N AA,Acute blood loss, HA, Macrocytic100 32 N megaloblastic anemia Microcytic80 26 N anemia of chronic disease Microcytes/80 26 14.9% means that there is increased variations in cell size above the normal amount (anisocytosis) A normal val

21、ue means that the RBC population is uniform in size . Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 25 Osmotic fragility test 红细胞渗透脆性试验红细胞渗透脆性试验 Osmotic fragility test measures the RBC sensitivity to a hypotonic solution of saline. Saline concentrations of 0 to 0.9% are incubated

22、with RBCs at room temperature and the percent of hemolysis is measured. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 26 Osmotic fragility test Reference range Hemolysis start at 0.42%-0.46% Saline. Hemolysis complete at 0.28%-0.34% Saline. Liu, Yan-Fang MD, Ph. D. First Teaching

23、Hospital ZhengZhou Univ. 27 Osmotic fragility test Clinical significance Increase: Hereditary Spherocytosis Hereditary Elliptocytosis They have a limited ability take up water in a hypotonic solution and will, therefore, lyse at a higher sodium concentration than normal RBCs Decrease: Thalassemia, I

24、DA Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 28 Acidified serum test (Hams test) 酸溶血试验 Acidified serum test (Hams test) is the definitive diagnostic test for PNH. In acidified serum, complement is activated by the alternate pathway, binds to RBCs, and lyses the abnormal RBCs f

25、ound in PNH. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 29 Hams Test - Principle Normal RBCNormal RBC + Patients Serum + Patients Serum PNH RBCPNH RBC Patients SerumPatients Serum No hemolysisNo hemolysis Ph6.6-6.8Ph6.6-6.8 37lh37lh hemolysishemolysis Ph6.6-6.8Ph6.6-6.8 37lh37l

26、h Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 30 Clinical Significances Negative: Normal individuals Positive: Paroxysmal Nocturnal Hemoglobinuria High specificity, Low sensitivity Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 31 Sucrose hemolysis test 糖水试验 Su

27、crose hemolysis test sucrose provides a low ionic strength that permits binding of complement to RBCs. In paroxysmal nocturnal hemoglobinuria (PNH), the RBCs are abnormally sensitive to this complement mediated hemolysis. This is used in screening for PNH. High sensitivity and low specificity. Liu,

28、Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 32 ( (四四) )含铁血黄素尿试验（含铁血黄素尿试验（RousRous试验）试验） 原原 理理: :铁离子铁离子+ +低铁氰化钾低铁氰化钾 低铁氰化铁低铁氰化铁( (蓝色）蓝色） 正常人正常人: :（- -） 意意 义义: :（+ +）说明慢性血管内溶血，）说明慢性血管内溶血， 常见于常见于PNHPNH Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 33 Coombs (Direct An

29、tiglobulin) test抗抗 人球蛋白试验人球蛋白试验 Critical in the evaluation of hemolytic anemia, especially when autoimmune etiology is suspected Anti-globulin reagent prepared by injecting human globulins into animals-antibody isolated IgG C3 (Marker for IgM) Mix Coombs reagent with patients RBC and test for clotti

30、ng in the test tube Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 34 Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 35 Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 36 Direct Coombs Test Human Globulin Antibodies to Human Globulins +Patient RBC

31、 Ab Ab Ab Ab Agglutination Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 37 Clinical Significances Negative: Normal individuals. Positive:AIHA Direct Coombs test: AIHA, Hemolysis in infants Indirect Coombs test: Rh or ABO not matched pregnancy. Others with positive results of Coom

32、bs test SLE, R.A., Lymphoma, Drug-related hemolysis Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 38 Hemoglobin electrophoresis 血红蛋白电泳 Hemoglobin electrophoresis can be used to identify the presence of an abnormal hemoglobin (called hemoglobinopathies). Different hgbs will move to

33、 different regions of the gel and the type of hemoglobin may be identified by its position on the gel after electrophoresis. Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 39 The lab investigation may also include A bone marrow smear and biopsy Used when other tests are not conclus

34、ive In a bone marrow sample, the following things should be noted: Maturation of RBC and WBC series Ratio of myeloid to erythroid series Abundance of iron stores (ringed sideroblasts) Presence or absence of granulomas, tumor cells Red to yellow ratio Presence of megakaryocytes Liu, Yan-Fang MD, Ph. D. First Teaching Hospital ZhengZhou Univ. 40 Enzyme assay Evaluation of RBC enzymes and metabolic pathways enzyme deficiencies in carbohydrate metabolic path