Hematology Lab - 09 Automation

HEMATOLOGY LABORATORY AUTOMATION AUTOMATION • • • • • • Most clinical laboratories uses automated analyzers for blood cell counting Automated analyzers can produce results in short period of time Manual methods o Take longer time to accomplish o Requires mastery of cell morphology (esp. differential ct.) Included parameters o Complete Blood Count ▪ RBC Count ▪ WBC Count ▪ Platelet Count ▪ Hemoglobin Count ▪ Hematocrit Count ▪ WBC Differential Count ▪ RBC Indices (MCV, MCH, MCHC) Most common principles used o Electrical Impedance (electrical impulses) o Optical Scatter (angle of light) Automation is continually developed o Equipment used ▪ Analog or digital ▪ Display features ▪ Barcode reader (for patient id) o Instrumental capabilities o Can it show flags, abnormalities, critical values? ▪ Critical values - reported to physician; • First, recheck sample when occurs • Then, rerun test with control solution • Report occurrence when critical value repeats o Data output ▪ Histograms, graphs ▪ Paperless recording ▪ Direct reporting of results to physician ELECTRICAL IMPEDANCE • • • • • • • • • • aka Low-Voltage Direct Current or Coulter Principle Developed by Coulter Electronics (1950’s) Coulter Cell Counter Model A o First automated hematology analyzer Most commonly employed principle in automated hematology analyzers PRINCIPLE • As a dilute suspension of cells is drawn through the aperture, the passage of each individual cell momentarily increases the impedance (resistance) of the electrical path between two submerged electrodes that are located on each side of the aperture o Cells are poor conductors of electricity o Measures electrical pulses produced as cell pas through the aperture o RBC aperture - measures RBC & plt simulateneously ▪ Hemolyzes RBC after measurement to prevent interference with WBC count and simultaneously measure hemoglobin o WBC aperture - for WBC count Number of pulses (y axis) = Number of cells Amplitude of pulse (x-axis) = Cell size/volume o Threshold circuit (allows discrimination of cells of specific volume) Plotted on a volume distribution histogram Errors which may be encountered o Instrumental Errors ▪ Aperture plugs - positive error (false ↑) ▪ Bubbles in the specimen - positive error (false ↑) ▪ Extraneous electrical impulses - pos error (false ↑) ▪ Excessive lysing of RBCs - negative error (false ↓) ▪ Improper setting of aperture current - pos/neg error o Nature of Specimen ▪ Giant platelets • Counted as RBC or WBC • False↑ - RBC, WBC; False↓ - plt ct ▪ Fragments of WBC cytoplasm - counted as plt or RBC • False↑ - RBC, plts ▪ Some abnormal RBCs resists lysis (False↑ - WBC ct) OPTICAL SCATTER • • Uses laser light (mercury arc) or non-laser light (tungsten halogen lamp) Used to measure light scattering properties of cells o Measure angles that are relevant specific characteristic of cells • PRINCIPLE • • • • • • • As a single cell passes through the sensing zone of flow cell, it scatters the focus light Scattered light is detected by a photodetector o Photodetector - converts scattered light to electrical pulses The number of pulse is directly proportional to the number of cells passing through the sensing zone. OPTICAL LIGHT SCATTER PROPERTIES • • • • Absorbance – wavelength of light that is not transmitted Diffraction – bending of light around corners with the use of small angles Refraction - bending of light because of a change in speed with the use of intermediate angles Reflection - light rays turned back by the surface or an obstruction with the use of large angles ANGLES OF LIGHT SCATTER • • • Forward-angle light scatter (0°) – cell volume or cell size o Result of the diffraction of light produced by cell Forward low-angle light scatter (2 to 3°) & Forward high-angle (5 to 15°) - can relate to size or volume and allows for description of the refractive index of cellular components Orthogonal light scatter (90°) aka side scatter correlates w/ degree of internal complexity (granularity and lobularity) o Allow identification of granulocytes o Result of reflection and refraction of light by internal cell components CRITICAL VALUES • Checking of sample when critical values are present o Rerun sample with control solution (to check accuracy of analyzer) o Manual method (to check result of analyzer) o Repetition of values after reruns and manual methods should be reported to physician TESTS CRITICAL VALUES Hematocrit ≤ 21% (0.21) ≥ 65% (0.65) Hemoglobin ≤ 70 g/L ≥ 200 g/L Reticulocyte > 20% WBC Count < 2,000/uL > 50,000/uL Blood Smear Neutrophilic phagocytosis of microorganism (systemic infection) Abnormal Leukemoid Reaction Schistocytes (fragmented RBC - hemolytic anemia) Sickle cells (hemoglobinopathies, blasts) Blasts (if new patient) (leukemic) Presence of intracellular organisms Platelets < 20,000/uL (if not previously reported) > 1 million/uL • Neutrophilic phagocytosis of microorganism - systemic infection Abnormal Leukemoid Reaction Schistocytes - fragmented RBC - hemolytic anemia Sickle cells - hemoglobinopathies, blast Blasts - may indicate leukemic reaction o New patient - reported immediately o Old patient - check record ▪ If no previous record, report immediately ▪ If previous records include blast, no need for immediate reporting as physician is informed Intracellular organisms (malaria, bacterial)