Urinalysis and Body Fluids CRg Student review for Competency and BF Lab Practical

Neubauer hemacytometer / counting chamber Large squares, such as # 1-9 below have volume of 0.1 Small squares – in center # 5 have volume of 0.004 RBC and WBC counts are reported as # cells / uL To obtain this you must Count and record cells from both sides of the chamber. Average the two sides And enter this information into the formula shown The average number of cells from both sides of the chamber are multiplied by the dilution factor . This number will be 1 ,unless you needed to make a dilution. ( On the denominator of the calculation, you enter the number of squares you counted each side of the chamber. Times the volume of each square. Large squares, such as # 1-9 seen on the picture below have volume of 0.1 Small squares – in center # 5 have volume of 0.004

Improved Neubauer hemacytometer / counting chamber Counting the cells Cells touching either the top or left boundary are included in the count When triple lines are present, the MIDDLE line is used as the boundary.

Neubauer hemacytometer / counting chamber Formula for calculations – results in # cells / uL Count and record cells from both sides of the chamber. Average the two sides Multiply by dilution factor (if no dilution is made, this number is 1) Divide by number of squares counted X volume of each square Large squares, such as # 1-9 below have volume of 0.1 Small squares – in center # 5 have volume of 0.004

Body Fluid Differential Specimens for cell differential – usually collected in EDTA tube (except CSF samples) Cytospin preparations produce more accurate counts, however cell distortion often seen (can be minimized by adding albumin to sample before processing). Stain = Wright/Wright-Giemsa (Gram stain may be used if bacterial infection suspected.) Hyaluronidase added to synovial fluid to reduce the viscosity and make the slide more uniform.

Body Fluid Differential Any nucleated peripheral blood cell, including immature forms, plasma, and LE cells can be found. Check textbook / classroom notes for normal values. Monocytes/macrophages with inclusions are renamed Erythrophages contain one or more red cells Siderophages contain hemosiderin /siderotic bodies or hematin crystals Leukophages contain white blood cell(s) Lipophage contains ingested fat/lipid

Body Fluid Differential Entire smear should be evaluated for abnormal cells inclusions within cells Clusters Presence of intracellular organisms

Body Fluid Differential CSF specimen cells L – neutrophiles & @6 macrophages R – neutrophiles and intracellular bacteria

Body Fluid Differential - CSF Eosinophils Often associated with parasitic / fungal infections, allergic reactions including reaction to shunts and other foreign objects.

Body Fluid Differential - CSF Ependymal cells Normal cell, unique to CSF Line the ventricles, produce CSF fluid Large cell with distinct round/oval nucleus, sometimes found in sheets

Body Fluid Differential - CSF Cellular inclusions Erythrophage Siderophage Hematoidin crystals (see below) ASCP 21 CSF erythrophage, with few iron granules forming 1991 CAP 30 CSF hematoidin crystal / bilirubin crystal ASCP 6 macrophage, lymphocyte, siderophage

Body Fluid Differential - CSF India-ink / nigrosin preparation Negative stain to view the encapsulated Cryptococcus neoformans (frequent complication seen in AIDs /immunocompromised patients) Instead of using stain, can also use dark field microscopy to get same effect. Has @ 25-50% sensitivity Serological latex agglutination tests provide better results – and are preferred.

Body Fluid Differential Suspicious / unclassified or malignant cells are reported as “other” or “unclassified” AND are sent to pathology. (as seen below) Cytology – send unstained slide to cytology / pathology 1986 CAP CM10 CSF – blasts (appearance similar to peripheral blood, always consult with hematology specialist / pathologist) ( see below right)

Body Fluid Differential left is 1988 CAP CM 25 – CSF 250x malignant cells Remember – we classify them as ‘other’ or ‘unclassified’ and take the slide to the cytologist / pathologist Right is leukemic cells found in CSF

Body Fluid Differential - Serous Serous fluid examples Left -lymphocyte, macrophage/mono, basophil Right - Mesothelial cells

Serous fluid - Mesothelial cell

Serous Fluids Malignant cells ACSP 7 Case 1 cont. peritoneal fluid, malignancy

Serous Fluids Malignant cells ASCP 12 Case 4, 30 year old with back pain and inability to work. Pleural effusion fluid – malignant tumor on spinal cord

Serous Fluids-LE cells Seen in patients with Systemic Lupus Erythmatosis (SLE) a systemic disease in which an autoantibody attacks the patients organs and body systems LE cell is a neutrophil or macrophage that has engulfed a homogeneous mass of purple staining nuclear material It is not unusual to find macrophages engulfing other cells,. If the nucleus of the ingested cell has intact nuclear structures / normal chromatin pattern, the macrophage is considered to be a Tart cell . Tart cells are not associated with SLE. Example is a segmented neutrophil and demonstrates the LE cell characteristic of containing a smooth homogenous nuclear mass.

Synovial Fluid In addition to counting and identifying cells; identification of any crystals present aids in diagnosis. Monosodium urate associated with gouty arthritis. Calcium pyrophosphate associated with pseudogout. Blunt ended rod-like crystals resembling monosodium urate, crystals can be seen in patients following joint steroid injection. Others: oxalate and cholesterol

The Gout 1799 caricature by James Gillray