ASCP SH cheat sheet

Hematopoiesis shifts from yolk sac (weeks 3-8) to liver and spleen (weeks 6 to birth) to bone marrow (from about week 20 onward); adult hematopoiesis occurs in the marrow of the axial skeleton.

Erythropoietin is produced by peritubular interstitial cells of the kidney in response to tissue hypoxia and drives CFU-E proliferation and differentiation into erythroblasts.

Adult reference values: Hb 13.5-17.5 g/dL (male) and 12.0-15.5 g/dL (female); Hct 41-53% (male) and 36-46% (female); RBC 4.5-5.9 and 4.1-5.1 x 10^12/L; MCV 80-100 fL; MCH 27-33 pg; MCHC 32-36 g/dL; RDW 11.5-14.5%.

Reticulocyte reference is 0.5-2.5%; corrected retic = retic% x (patient Hct / 45); RPI = corrected retic / maturation factor (1.0 at Hct 45, 1.5 at 35, 2.0 at 25, 2.5 at 20).

Adult hemoglobin: HbA (alpha2 beta2) about 95-97%, HbA2 (alpha2 delta2) below 3.5%, HbF (alpha2 gamma2) below 2%; each heme group is protoporphyrin IX plus ferrous iron.

Right shift of the oxygen dissociation curve (decreased affinity, increased tissue delivery) occurs with increased 2,3-BPG, hydrogen ion, carbon dioxide, and temperature; normal P50 is about 26-27 mmHg.

Red cell metabolic routes: glycolysis generates ATP, the hexose monophosphate shunt generates NADPH, the Rapoport-Luebering shunt generates 2,3-BPG, and cytochrome b5 reductase reduces methemoglobin.

Membrane skeleton: defective vertical links (spectrin-ankyrin-band 3, protein 4.2) cause hereditary spherocytosis; defective horizontal links (spectrin dimer contacts, protein 4.1) cause hereditary elliptocytosis.

Iron balance: total body iron 3-5 g; duodenal absorption 1-2 mg/day; transferrin transports iron, ferritin and hemosiderin store it, and hepcidin inhibits absorption and macrophage iron release.

Anemia is a hemoglobin below the age- and sex-matched reference interval; morphologic classification is microcytic (MCV below 80 fL), normocytic (80-100 fL), or macrocytic (above 100 fL).

Iron deficiency progresses from storage depletion (low ferritin) to iron-deficient erythropoiesis (low saturation, high soluble transferrin receptor, high zinc protoporphyrin) to anemia; ferritin below 15 ng/mL is highly specific.

Anemia of chronic disease: hepcidin-mediated iron sequestration gives low serum iron, low TIBC, normal or elevated ferritin, and saturation usually 15-20%; the anemia is normocytic and normochromic.

Beta thalassemia trait: HbA2 3.5-8% with microcytosis and an increased RBC count, normal iron studies; beta thalassemia major shows severe anemia, prominent HbF, and extramedullary hematopoiesis.

Alpha thalassemia by number of deleted genes: one is a silent carrier, two is trait, three is HbH disease (beta4 tetramers), and four is Hb Bart hydrops fetalis.

Sickle cell disease: beta6 glutamic acid replaced by valine; HbS migrates more slowly than HbA on alkaline electrophoresis; the solubility test is screening only, confirmed by HPLC, electrophoresis, or DNA testing.

Megaloblastic anemia: elevated MCV, hypersegmented neutrophils, high LDH and indirect bilirubin, low haptoglobin; B12 deficiency raises methylmalonic acid and homocysteine, folate deficiency raises homocysteine alone.

Hemolysis markers: increased reticulocytes, indirect bilirubin, and LDH with decreased haptoglobin; the direct antiglobulin test separates immune (positive) from non-immune causes.

Hereditary spherocytosis: spherocytes, increased MCHC and RDW, increased osmotic fragility, reduced eosin-5-maleimide binding, negative DAT; paroxysmal nocturnal hemoglobinuria shows CD55/CD59-deficient clones by flow cytometry.

Adult WBC reference is 4.5-11.0 x 10^9/L with neutrophils 40-70%, bands 0-5%, lymphocytes 20-45%, monocytes 2-10%, eosinophils 1-4%, and basophils 0-2%.

Leukemoid reaction: left shift with toxic granulation and Dohle bodies, high leukocyte alkaline phosphatase, no basophilia; CML shows basophilia, low LAP, and t(9;22) BCR-ABL1.

AML: myeloblasts with Auer rods, positive myeloperoxidase and Sudan black B, negative TdT, and myeloid markers CD13, CD33, CD117.

Acute promyelocytic leukemia (FAB M3): t(15;17) PML-RARA, hypergranular promyelocytes, DIC at presentation; all-trans retinoic acid plus arsenic trioxide is targeted therapy.

ALL: TdT-positive blasts; B-ALL expresses CD19, CD10, CD22; T-ALL expresses CD3 and CD7; peak age 2-5 years and central nervous system prophylaxis is required.

CLL: absolute lymphocytosis above 5 x 10^9/L, smudge cells on the film, CD5+, CD19+, CD23+, dim surface immunoglobulin, FMC7 negative.

Hairy cell leukemia: cells positive for CD11c, CD25, CD103, and tartrate-resistant acid phosphatase; pancytopenia with monocytopenia, splenomegaly, dry marrow tap, and fried-egg marrow cells.

Classic Hodgkin lymphoma: Reed-Sternberg cells are CD15+, CD30+, CD45-, usually CD20-; nodular lymphocyte predominant Hodgkin lymphoma is CD20+ and CD15/CD30 negative.

Multiple myeloma: CD38+/CD138+ plasma cells, monoclonal M-spike, rouleaux, CRAB features (hypercalcemia, renal failure, anemia, bone lesions) and urinary Bence-Jones protein.

Primary hemostasis: vascular constriction, platelet adhesion through glycoprotein Ib and von Willebrand factor, then activation and aggregation through glycoprotein IIb/IIIa and fibrinogen.

Intrinsic factors are XII, XI, IX, VIII; extrinsic is factor VII with tissue factor; common pathway is X, V, II, I; APTT screens intrinsic plus common, PT screens extrinsic plus common.

Reference intervals: platelets 150-450 x 10^9/L, PT 11-15 s, INR 0.8-1.2, APTT 25-35 s, fibrinogen 200-400 mg/dL, thrombin time 15-20 s, D-dimer below 500 ng/mL FEU.

Mixing study: immediate correction indicates factor deficiency; failure to correct immediately or after 1-2 hours at 37 C indicates a specific inhibitor, a lupus anticoagulant, or heparin contamination.

von Willebrand disease: low von Willebrand factor antigen, low ristocetin cofactor activity, low factor VIII; abnormal ristocetin aggregation that corrects with normal plasma.

Hemophilia A (factor VIII) and B (factor IX) are X-linked; APTT is prolonged, PT and platelet count are normal, mixing corrects, and inhibitors are quantitated by Bethesda assay.

Vitamin K deficiency reduces factors II, VII, IX, X and proteins C and S; PT prolongs before APTT because factor VII has the shortest half-life.

DIC: thrombocytopenia, prolonged PT and APTT, low fibrinogen, elevated D-dimer and fibrin degradation products, schistocytes; serial testing is more informative than a single set.

Anticoagulant monitoring: unfractionated heparin by APTT or anti-Xa and reversed by protamine; low molecular weight heparin by anti-Xa; warfarin by PT/INR and reversed by vitamin K and factor concentrates.

Specimen anticoagulants: EDTA (K2 or K3) for CBC and morphology; 3.2% sodium citrate at 9 parts blood to 1 part anticoagulant for PT and APTT; lithium heparin for chemistry and selected osmotic fragility studies.

Stability: EDTA CBC samples are stable about 4-8 hours at room temperature for morphology; PT and APTT should be performed within 4 hours on capped or separated plasma, or plasma frozen at -20 C or colder.

Impedance counters size and count cells by electrical resistance; optical light scatter and flow cytometry using volume, conductivity, and scatter generate five-part leukocyte differentials.

Cold agglutinin interference: falsely low RBC and platelet counts with falsely high MCV and MCHC; warm the EDTA specimen to 37 C, mix, and rerun before reporting.

Westgard multirule: 1-2s is a warning only; reject the run on 1-3s, 2-2s, R-4s, 4-1s, and 10x rule violations.

Calibration establishes the measuring relationship, controls monitor stability, Levey-Jennings charts reveal trends and shifts, and delta checks compare a current result with the patient's previous values.

ESR by Westergren is elevated by anemia, pregnancy, and inflammation and reduced by polycythemia, spherocytosis, and sickle cells; it is a nonspecific acute-phase indicator.

Reticulocytes require supravital stains (new methylene blue, brilliant cresyl blue) that precipitate ribosomal RNA, or flow cytometry with fluorescent nucleic acid dyes.

Quality requirements include documented proficiency testing, QC before patient testing, reference interval verification, linearity checks, corrective action records, and critical value reporting.

Specimen anticoagulants: EDTA (K2 or K3) for CBC and morphology; 3.2% sodium citrate at 9 parts blood to 1 part anticoagulant for PT and APTT; lithium heparin for chemistry and selected osmotic fragility studies.

Stability: EDTA CBC samples are stable about 4-8 hours at room temperature for morphology; PT and APTT should be performed within 4 hours on capped or separated plasma, or plasma frozen at -20 C or colder.

Impedance counters size and count cells by electrical resistance; optical light scatter and flow cytometry using volume, conductivity, and scatter generate five-part leukocyte differentials.

Cold agglutinin interference: falsely low RBC and platelet counts with falsely high MCV and MCHC; warm the EDTA specimen to 37 C, mix, and rerun before reporting.

Westgard multirule: 1-2s is a warning only; reject the run on 1-3s, 2-2s, R-4s, 4-1s, and 10x rule violations.

Calibration establishes the measuring relationship, controls monitor stability, Levey-Jennings charts reveal trends and shifts, and delta checks compare a current result with the patient's previous values.

ESR by Westergren is elevated by anemia, pregnancy, and inflammation and reduced by polycythemia, spherocytosis, and sickle cells; it is a nonspecific acute-phase indicator.

Reticulocytes require supravital stains (new methylene blue, brilliant cresyl blue) that precipitate ribosomal RNA, or flow cytometry with fluorescent nucleic acid dyes.