Template:Clinical pathology - entire topic

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Clinical pathology

Author: Mikael Häggström, M.D. [note 1]
Memorization-worthy:[note 2] For the most likely types of cases and/or questions that you may be responsible for, know where to find local policies and procedures, and have a good idea of whom to ask for further advice. Make sure you have access and/or contact details for whenever and wherever you are likely to need them.


Sample tube types

The following is an overview of the main types of sample tube types, organized by order of draw (the recommended sequence by which fluid is drawn) during blood draws:

Tube cap color or type in order of draw Additive Usage and comments
Blood culture bottle Sodium polyanethol sulfonate (anticoagulant) and growth media for microorganisms Usually drawn first for minimal risk of contamination.[1] Two bottles are typically collected in one blood draw; one for aerobic organisms and one for anaerobic organisms.[2]
Light blue Sodium citrate (anticoagulant) Coagulation tests such as prothrombin time (PT) and partial thromboplastin time (PTT) and thrombin time (TT). Tube must be filled 100%.
Plain red No additive Serum: Total complement activity, cryoglobulins
Gold (sometimes red and grey "tiger top"[3]) Clot activator and serum separating gel[4] Serum-separating tube: Tube inversions promote clotting. Most chemistry, endocrine and serology tests, including hepatitis and HIV.
Dark green Sodium heparin (anticoagulant) Chromosome testing, HLA typing, ammonia, lactate
Light green Lithium heparin (anticoagulant) Plasma. Tube inversions prevent clotting
Lavender ("purple") EDTA (chelator / anticoagulant) Whole blood: CBC, ESR, Coombs test, platelet antibodies, flow cytometry, blood levels of tacrolimus and cyclosporin
Pink EDTA (chelator / anticoagulant) Blood typing and cross-matching, direct Coombs test, HIV viral load
Royal blue EDTA (chelator / anticoagulant) Trace elements, heavy metals, most drug levels, toxicology
Tan EDTA (chelator / anticoagulant) Lead
Gray
  • Sodium fluoride (glycolysis inhibitor)
  • Potassium oxalate (anticoagulant)[5]
Glucose, lactate[6]
Yellow Acid-citrate-dextrose A (anticoagulant) Tissue typing, DNA studies, HIV cultures
Pearl ("white") Separating gel and (K2)EDTA PCR for adenovirus, toxoplasma and HHV-6

Lab management

Lab management is essentially about handling each of the extremely various lab-related situations that arise, and can generally be achieved by:

  • Common sense
  • Gathering enough information before a decision
  • Identifying what questions needs answering
  • Asking proper expertise and/or looking up relevant information in proper sources (see learning pathology), which may include local protocols as well as policies of accrediting organizations of the department (which are generally more stringent than the national or regional laws).

Test question

Retention time

(You may skip this question if you don't expect to ever be part of laboratory management in the US.)

You work in a pathology department in the United States, which is accredited by the College of American Pathologists (CAP). Your local procedure manual states that non-forensic paraffin-embedded blocks must be retained for at least 10 years before being thrown away. In order to save storage space, one suggestion that gets brought up is to reduce the retention time to 5 years. You look up the issue, and find that federal U.S. law (the Clinical Laboratory Improvement Amendments; CLIA) states that such blocks must be retained for at least 2 years. Is it acceptable to finish the look-up here, and agree to reduce the retention time of non-forensic paraffin-embedded blocks to 5 years in this department?

  • Answer: This pathology department is accredited by CAP, whose requirements commonly exceed those of CLIA, in this case stating at least 10 years for non-forensic paraffin-embedded blocks. Therefore, it is not acceptable to relax minimum retention times without first having had a look at CAP requirements, if the lab is accredited by it (don't assume it isn't without checking). Thus, the answer to this question is: no. Furthermore, state laws need to be considered as well, as they may exceed those of CLIA.

For quick look-up in the future, the following are the most relevant retention times, as given by CLIA[7] as well as by CAP[8] (more comprehensive lists are available in their sources):

Microscopy slides Histology and non-forensic autopsy 10 years[7]
Forensic autopsy Indefinitely[7]
Cytology, fine needle aspiration 10 years[8]
Cytology, apart from fine needle aspiration 5 years[7]
Paraffin-embedded blocks Non-forensic 2[7] or 10 years[8]
Forensic Indefinitely[7]
Requisition forms and test reports Pathology reports 10 years[7]
Other 2 years[7]
Blood bank records Quality control records 5 years[8]
Donor and recipient records 10 years[8]
Records of indefinitely deferred donors Indefinitely[8]
Wet tissues Until report is completed[7] or 2 weeks thereafter[8]
Proficiency testing records and quality management/quality control records (except for blood bank) 2 years[7]
Discontinued procedures 2 years[7]
Blood smears and other body fluid smears, microbiology slides (including Gram stains) 7 days[8]
Flow cytometry plots 10 years[8]

Notes

  1. For a full list of contributors, see article history. Creators of images are attributed at the image description pages, seen by clicking on the images. See Patholines:Authorship for details.
  2. Further information on what is memorization-worthy or not: Learning pathology

Main page

References

  1. Pagana, KD; Pagana, TJ; Pagana, TN (19 September 2014). Mosby's Diagnostic and Laboratory Test Reference - E-Book . Elsevier Health Sciences. ISBN 978-0-323-22592-2. 
  2. "Chapter 3.4.1: Blood cultures; general detection and interpretation". Clinical Microbiology Procedures Handbook . Wiley. 6 August 2020. ISBN 978-1-55581-881-4. 
  3. . Test Tube Guide and Order of Draw. Guthrie Laboratory Services (June 2019).
  4. . Specimen requirements/containers. Pathology & Laboratory Medicine, UCI School of Medicine.
  5. "Effects of standard anticoagulants and storage procedures on plasma glucose values in seals. ". J Am Vet Med Assoc 201 (1): 145–8. 1992. PMID 1644639. Archived from the original. . 
  6. Amitava Dasgupta; Jorge L. Sepulveda (20 July 2019). Accurate Results in the Clinical Laboratory: A Guide to Error Detection and Correction . Elsevier Science. p. 131. ISBN 978-0-12-813777-2. 
  7. 7.00 7.01 7.02 7.03 7.04 7.05 7.06 7.07 7.08 7.09 7.10 . 42 CFR § 493.1105 - Standard: Retention requirements.. Cornell Law School. [68 FR 3703, Jan. 24, 2003; 68 FR 50723, Aug. 22, 2003]
  8. 8.0 8.1 8.2 8.3 8.4 8.5 8.6 8.7 8.8 . CAP Policy Manual - Policy PP. Minimum Period of Retention of Laboratory Records and Materials. CAP.org. Adopted August 1995. Revised September 2020

Image sources


Blood bank

Author: Mikael Häggström [note 1]
If you expect to get questions regarding blood products, get a copy of the local cutoffs for approving transfusions of red blood cells, platelets and plasma, and keep it so that you can quickly look it up when needed.

Always consider if a blood product should be given as per a local precaution protocol (usually including regular check-ups of the patient).

Specific questions or requests

  • Generalized dosage in adults:
  • Plasma: Generally 10-15 ml/kg, and results in approximately 25-30% plasma volume replacement. Detailed calculation
  • Cryoprecipitate: 1 unit for every 7-10kg of body weight. 10 units generally replaces 100-150 mg/dl. Detailed calculation

Notes

  1. For a full list of contributors, see article history. Creators of images are attributed at the image description pages, seen by clicking on the images. See Patholines:Authorship for details.

Main page

References


Image sources


Blood compatibility testing

Author: Mikael Häggström [note 1]

Blood typing

ABO antigens and antibodies

Upon direct testing by adding antibodies against A, B and/or Rh to patient blood, agglutination means that the patient has the antigen tested. Upon indirect testing by adding A or B antigen to patient plasma, agglutination means absence of the antigen in the patient (and thus the patient produces antibodies against it).

Other blood group systems

In the antibody screening procedure, an individual's plasma is added to a panel of two or three sets of red blood cells which have been chosen to express most clinically significant blood group antigens. Agglutination of the screening cells by the plasma, with or without the addition of anti-human globulin, indicates that an unexpected blood group antibody is present. If this occurs, further testing using more cells (usually 10–11) is necessary to identify the antibody. By examining the antigen profiles of the red blood cells the person's plasma reacts with, it is possible to determine the antibody's identity.

The image above shows the interpretation of an antibody panel used to detect antibodies towards the most relevant blood group antigens. Each row represents "reference" or "control" red blood cells of donors which have known antigen compositions and are ABO group O. A + means that the antigen is present on the reference red blood cells, and 0 means it is absent; nt means "not tested". The "result" column to the right displays reactivity when mixing reference red blood cells with plasma from the patient in 3 different phases: room temperature, 37°C and AHG (with anti-human globulin, by the indirect antiglobulin test).[1]

  • Step 1; Annotated in blue: starting to exclude antigens without reaction in all 3 phases; looking at the first reference cell row with no reaction (0 in column at right, in this case cell donor 2), and excluding (here marked by X) each present antigen where the other pair is either practically non-existent (such as for D) or 0 (presence is homozygous, in this case homozygous c).
    When both pairs are + (heterozygous cases), they are both excluded (here marked by X), except for C/c, E/e, Duffy, Kidd and MNS antigens (where antibodies of the patient may still react towards blood cells with homozygous antigen expression, because homozygous expression results in a higher dosage of the antigen).[2] Thus, in this case, E/e is not excluded in this row, while K/k is, as well as Jsb (regardless of what Jsa would have shown).[note 2]
  • Step 2: Annotated in brown: Going to the next reference cell row with a negative reaction (in this case cell donor 4), and repeating for each antigen type that is not already excluded.
  • Step 3: Annotated in purple. Repeating the same for each reference cell row with negative reaction.
  • Step 4: Discounting antigens that were absent in all or almost all reactive cases (here marked with \). These are often antigens with low prevalence, and while there is a possibility of such antibodies being produced, they are generally not the type that is responsible for the reactivity at hand.
  • Step 5: Comparing the remaining possible antigens for a most likely culprit (in this case Fya), and selectively ruling out significant differential antigens, such as with the shown additional donor cell type that is known to not contain Fya but contains C and Jka.

In this case, the antibody panel shows that anti-Fya antibodies are present. This indicates that donor blood typed to be negative for the Fya antigen must be used. Still, if a subsequent cross-matching shows reactivity, additional testing should be done against previously discounted antigens (in this case potentially E, K, Kpa and/or Lua).[1]

Hemagglutination inhibition[2]
Neutralizing substance Antigen cancelled
  • Hydatid cyst fluid
  • Pigeon eggs
P1
Saliva H, Lea
Breast milk I
Guinea pig urine Sda

When multiple antibodies are present, or when an antibody is directed against a high-frequency antigen, the normal antibody panel procedure may not provide a conclusive identification. In these cases, hemagglutination inhibition can be used, wherein a neutralizing substance cancels out a specific antigen.[2] Alternatively, the plasma may be incubated with cells of known antigen profiles in order to remove a specific antibody (a process termed adsorption); or the cells can be treated with enzymes such as ficain or papain which inhibit the reactivity of some blood group antibodies and enhance others (see table below).

People who have tested positive for an unexpected blood group antibody in the past may not exhibit a positive reaction on subsequent testing; however, if the antibody is clinically significant, they must be transfused with antigen-negative blood regardless.[3]

Following is a comparison of clinically relevant characteristics of antibodies against the main human blood group systems:[2]

ABO Rh Kell Duffy Kidd Lutheran MNS Lewis P Ii
Most common in immediate hemolytic transfusion reactions A Yes Fya Jka
Most common in delayed hemolytic transfusion reactions E,D,C Jka
Most common in hemolytic disease of the newborn Yes D,C Yes
Commonly produce intravascular hemolysis Yes Yes Yes
Reactive at room temperature Yes M,N Lea, Leb P1
Nearly always clinically insignificant Yes M,N Yes P1
Naturally occurring Yes Yes M,N Yes Yes Yes
Enhanced by ficain[4] and papain[5] Yes Yes Yes Yes P1 Yes
Destroyed by ficain[4] and papain[5] Fya, Fyb Yes Yes
Displaying dosage Further information: Blood compatibility testing Cc, Ee Yes Yes Yes

Notes

  1. For a full list of contributors, see article history. Creators of images are attributed at the image description pages, seen by clicking on the images. See Patholines:Authorship for details.
  2. Besides from C/c, E/e, Duffy, Kidd and MNS, clinically significant dosage effects is rare but not impossible for other antigens, which thus may still be considered if subsequent cross-matching is reactive.

Main page

References

  1. 1.0 1.1 Justin R. Rhees, M.S., MLS(ASCP)CM, SBBCM. Introduction to Antibody Identification. University of Utah, Medical Laboratory Sciences.
  2. 2.0 2.1 2.2 2.3 Mais, Daniel (2014). Quick compendium of clinical pathology . United States: American Society for Clinical Pathology Press. ISBN 978-0-89189-615-9. OCLC 895712380. 
  3. Uhl, L (11 Jan 2021). Pretransfusion testing for red blood cell transfusion. UpToDate.
  4. 4.0 4.1 Hill, Ben C.; Hanna, Courtney A.; Adamski, Jill; Pham, Huy P.; Marques, Marisa B.; Williams, Lance A. (2017). "Ficin-Treated Red Cells Help Identify Clinically Significant Alloantibodies Masked as Reactions of Undetermined Specificity in Gel Microtubes ". Laboratory Medicine 48 (1): 24–28. doi:10.1093/labmed/lmw062. ISSN 0007-5027. PMID 28007780. 
  5. 5.0 5.1 Eric Ching. Questions and Answers on Proteolytic Enzymes Used in Blood Group Serology. Canadian Society for Transfusion Medicine.

Image sources


Kleihauer–Betke test

Author: Mikael Häggström [note 1]
The Kleihauer–Betke test stains fetal red blood cells (cells containing HbF) dark reddish-pink, while adult red blood cells will be white to light pink.

Collection

EDTA-containing-tube.

Criteria for fetal cells

KB stain with green marks at cells counted as fetal (HbF) cells, and red marks at incompletely colored cells at top and a too small cell at right.

To count, a fetal blood cell should be:

  • Stained more than approximately half of what is seen in control.
  • Not be nucleated or too big (white blood cell are generally also stained).
  • Not be too small.

Semi-quantification

This is done for Rh-negative mothers with a suspected feto-maternal hemorrhage, which can be suspected by various clinical findings (including neonatal anemia, stillbirth, intrauterine growth restriction, hydrops fetalis, decreased or absent fetal movements, non-reassuring fetal heart rate tracing, sinusoidal fetal tracing, and fetal tachyarrythmias, placenta previa with bleeding, and placental abruption)[1].

10HPFs (in 40x) are scanned, and fetal RBCs are counted (cells per 10 HPFs, not average per HPF), and classified as:

  • 0 - Negative
  • 1 - Rare
  • 2-5 - Few
  • 6-10 - Moderate
  • >10 - Abundant

Quantification

This is done for Rh-negative mothers to estimate the number of Rho(D) immune globulin vials to administer.

2000 cells are counted, in order to give a percentage calculated as:

  • Fetal RBCs (given in%) = (Fetal RBC count) / (Total cell count) *100

Alternatively, an acceptable estimation of at least 2000 cells can be done by using a micrograph (or a microscopy grid) to estimate the mean number of cells in a certain area, and using the same mean to estimate the number of cells in equally sized areas:
1. Count cells (both adult and fetal) until reaching 100 cells (including each cell in square by square if using a microscopy grid). Take note of how large micrograph area (or how many grid squares) were counted (here designated as x amount), and how many fetal RBCs were counted.
2. Pick another random location (you may randomize again if it is of a significantly different cell density, but do not let your decision be influenced by the number of fetal RBCs in the area or near its edge). Count the total number of cells in the same area size (or same x number of squares), and how many of them are fetal RBCs.
3. Pick another location again, and count the number of cells in the same area size, and how many of them are fetal RBCs.

  • If a count of 200-300 cells only shows 0 or 1 fetal RBC, there only needs to be 1 vial of 300 micrograms Rho(D) immune globulin, and the rest of the steps in this section can be skipped.
Standard
deviation
Count cells
in following
number of areas
Up to 6 3
7 4
8 5
9 6-7
10 8
11 10
12 12
  • Calculate how much the count for the second and third areas deviated from 100, and take the average thereof, which will be used as standard deviation.[note 2] If the standard deviation is higher than 12, count a total of 2000 cells regardless of areas and calculate as per formula above, and the rest of the steps can be skipped.
  • Use the table at right to estimate how many areas in total you need to count in order to have a mean number of cells per area with an acceptable confidence interval.

4. After having counted the needed number of areas, calculate the average of the number of cells per area (or per x number of squares), and assume that number for the rest of the counting.
5. Divide 2000 by the number of cells per area, and round that up to know the number of areas you need to perform the next step on in order to presumably have a total of 2000 cells (including previously counted areas).
6. In those additional areas, only count the number of fetal cells per area (or x number of squares), and add that to the fetal cells from previous areas.

Fetal RBCs (given in%) = (Fetal RBC count) / (Presumable total cell count) *100

Example:

The sum of all fetal RBCs in all areas in this example is 45. The presumable total cell count is 103 * 20 = 2060. Thus:

  • Fetal RBCs (given in%) = 45 / 2060 * 100 = 2.2%

Calculation of number of vials

Assuming that a vial of 300 micrograms of Rho(D) immune globulin will protect against 30 mL of fetal blood, the number of vials needed to compensate for the fetal-maternal transfusion is calculated as following, rounded up,[2] or rounded to the closest full number and then adding 1.[3]

Number of vials = Fetal RBCs in% * 1.7

For example, with 2.2% fetal RBCs, the number of vials would be 4[2] or 5[3].

Notes

  1. For a full list of contributors, see article history. Creators of images are attributed at the image description pages, seen by clicking on the images. See Patholines:Authorship for details.
  2. 2.0 2.1 Technically, the standard deviation would be calculated as the average deviation from the mean of all areas, but the simplified calculation used in this resource can be regarded to be close enough for practical purposes.

Main page

References

  1. Solomonia N, Playforth K, Reynolds EW (2012). "Fetal-maternal hemorrhage: a case and literature review. ". AJP Rep 2 (1): 7-14. doi:10.1055/s-0031-1296028. PMID 23946896. PMC: 3653511. Archived from the original. . 
  2. 2.0 2.1 Diann M. Krywko. Kleihauer Betke Test. StatPearls, National Center for Biotechnology Information. Last update: Last Update: January 20, 2020.
  3. 3.0 3.1 Practice at Danbury Hospital, Danbury, Connecticut, New England.

Image sources


Thromboelastography

Author: Mikael Häggström, M.D. [note 1]

Interpretation

Normal thromboelastogram with parameters.

Parameters derived from thromboelastography are mainly:[1]

  • R time: Time to initial clot formation (that is, amplitude deviation from baseline)
  • K time: Time from initial clot formation until reaching 20 mm in amplitude
  • Alpha angle (α): Angle between the baseline at initial clot formation, and a tangent line that intersects the tracing curve.
  • Maximum amplitude (MA): Maximum deviation of tracing to baseline.
  • A30: Amplitude 30 minutes after reaching maximum amplitude.

Following are examples of thromboelastography patterns and recommended treatments.[2][3]

Condition Appearance Main treatment
Normal Normal thromboelastography.png
Hemodilution or clotting factor deficiency Thromboelastography in hemodilution or clotting factor deficiency.png Fresh frozen plasma
Fibrinogen deficiency Thromboelastography in fibrinogen deficiency.png Cryoprecipitate
Low or dysfunctional platelets Thromboelastography in low or dysfunctional platelets.png Platelets
Thrombosis Thromboelastography in thrombosis.png Anticoagulant
Primary fibrinolysis Thromboelastography in primary fibrinolysis.png Antifibrinolytics or tranexamic acid
Secondary fibrinolysis Thromboelastography in secondary fibrinolysis.png Treating disseminated intravascular coagulation

Notes

  1. For a full list of contributors, see article history. Creators of images are attributed at the image description pages, seen by clicking on the images. See Patholines:Authorship for details.

Main page

References

  1. Tyler PD, Yang LM, Snider SB, Lerner AB, Aird WC, Shapiro NI (2021). "New Uses for Thromboelastography and Other Forms of Viscoelastic Monitoring in the Emergency Department: A Narrative Review. ". Ann Emerg Med 77 (3): 357–366. doi:10.1016/j.annemergmed.2020.07.026. PMID 32988649. 
  2. Collins S, MacIntyre C, Hewer I (2016). "Thromboelastography: Clinical Application, Interpretation, and Transfusion Management. ". AANA J 84 (2): 129–34. PMID 27311154. Archived from the original. . 
  3. Kreitzer NP, Bonomo J, Kanter D, Zammit C (2015). "Review of Thromboelastography in Neurocritical Care. ". Neurocrit Care 23 (3): 427–33. doi:10.1007/s12028-015-0187-9. PMID 26275677. Archived from the original. . 

Image sources


Peripheral blood smear

Author: Mikael Häggström [note 1]
Look at and comment separately on white blood cells, red blood cells and platelets. You generally don't need to aim for a perfect report, because for most purposes, a peripheral smear is a relatively simple screening test, and clinicians may opt to perform for example flow cytometry and/or a bone marrow biopsy if they want a better evaluation.

Comprehensiveness

On this resource, the following formatting is used for comprehensiveness:

  • Minimal depth
  • (Moderate depth)
  • ((Comprehensive))
Other legend

<< Decision needed between alternatives separated by / signs >>
{{Common findings / In case of findings}}
[[Comments]]
Link to another page

Oil immersion microscopy

This is preferred for light microscopy of peripheral blood smears in order to achieve a very high magnification. First use low or medium power to center on suspicious cells, or where red or white blood cells are best appreciated. Put a drop of immersion oil on the location and switch to the immersion objective. Then, whenever there's oil on a slide, always think twice before switching between objectives so as to avoid getting oil on any of your dry objectives (which is a bit tedious to clean off).

Red blood cells

Automated values

When available, automatic quantification of mean corpuscular volume (MCV) and red blood cell (RBC) distribution width (RDW), usually as part of CBC panel, generally decides whether you will call the sample "normocytic" versus "microcytic"/"macrocytic" and/or "anisocytotic", even if it is not clearly visible in the microscope. If automated values are not available, compare RBC sizes to lymphocyte nuclei, which should normally be the same size. If mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration (MCHC) are normal, but you still see multiple RBCs with central pallor greater than 50% of the diameter, you can report it as "Increased central pallor", and you may add "indicating iron deficiency" if it is compatible with the clinical history.

Automated values can be graded as follows:[1]

Interpretation Mild Moderate Marked
Microcytosis MCV : 70 - 79 MCV : 60 - 69 MCV <60
Macrocytosis MCV : 100 - 115 MCV : 115 - 125 MCV >125
Hypochromasia MCH : 23 - 26 MCH : 21 - 23 MCH <20
Anisocytosis RDW: 14.5[2] or 16[1] - 18 RDW : 18 - 22[1] or 26[2] RDW > 22[1] or 26[2]

Morphologic findings

Look for poikilocytosis (red blood cells of abnormal shapes). These are counted as a percentage of visible red blood cells:[1]

Image Rare/Occasional Moderate amount of Many/Abundant
Polychromasia Peripheral blood smear with polychromasia.jpg 3 - 5% 5 - 25% >25%
Spherocytes Micrograph of a spherocyte.jpg 1 - 5% 5 - 25% >25%
Schistocytes Micrograph of schistocytes.jpg up to 2% 2 - 25% >25%
Target cells (codocytes) Micrograph of a target cell.jpg up to 3% 3 - 25% >25%
Tear drop cells Micrograph of a tear drop cell (dacrocyte).jpg up to 2% 2 - 25% >25%
Burr cells (echinocytes) Micrograph of an echinocyte on a peripheral blood smear.jpg 1 - 3% 3 - 10% >10%
Sickle cells (drepanocytes) Micrograph of a sickle cell.jpg 3 - 5% 5 - 25% >25%
Elliptocytes Micrograph of an elliptocyte on a peripheral blood smear 02.jpg 1 - 5% 5 - 25% >25%
Basophilic stipplings Micrograph of a red blood cell with basophilic stippling.jpg up to 2% 2 - 25% >25%
Howell Jolly bodies Micrograph of a Howell–Jolly body in a red blood cell.jpg up to 1% 2 - 3 % >3%
Burr cells versus spur cells

Burr cells are distinguished from spur cells by having more equally distributed and rounded projections. They are usually artifactual. However, they may also be caused by renal insufficiency, so if this is present, a report may include "Occasional/Multiple echinocytes, consistent with renal insufficiency".

Intraerythrocytic findings

Platelets

If CBC is performed, use count to determine whether platelets are "normal in number" or whether there is "thrombocytopenia" or "thrombocytosis". If no CBC, count platelets within a high power oil immersed field, which should normally be 8 to 20.

A giant platelet.

Large platelets are those with a diameter greater than 4 microns. Giant platelets are those with a diameter greater than 7 microns (larger than a normal red blood cell).[3] Example report:

Numerous large and giant platelets(, suggesting an increased platelet turnover)(( such as in immune thrombocytopenic purpura. They may also be present in myeloproliferative neoplasms, myelodysplasia, and some congenital thrombocytopenia syndromes, including Bernard-Soulier syndrome and MYH9-related disorders.[3]))

In thrombocytopenia from automatic counting, look in particular for:

  • Clumping of platelets (which can cause a falsely low automatic platelet count). If present, check with the lab if it was sent in EDTA (which may cause artefactual clumping) and ask to have a blood sample sent in sodium citrate instead. Also, look for satellitosis (platelets attached around white blood cells).
  • Schistocytes among red blood cells.

White blood cells

Comparison of monoblast, promonocyte and monocyte. Further information: Suspected blasts on peripheral blood smear

Look for:

Report

Example report:

Normochromic normocytic red blood cells. Red blood cells show <normal morphology / anisopoikilocytosis with occasional ___>. [[If thrombocytopenia, also add "Schistocytes are not significantly increased" if applicable.]]

{{Leukocytosis with neutrophilia / lymphocytosis.}} White blood cells show no left shift or blasts.

Platelets show no evidence of clumping, and show normal granularity.

(Causes of the above findings include ___.)

Notes

  1. For a full list of contributors, see article history. Creators of images are attributed at the image description pages, seen by clicking on the images. See Patholines:Authorship for details.

Main page

References

  1. 1.0 1.1 1.2 1.3 1.4 Unless otherwise specified in table, reference is:
    - . Hong Kong Medical Technology Association - Quality Assurance Programme - Haematology and Serology, Prepared by HKMTAQAP Haematology & Serology Panel on November 2002..
  2. 2.0 2.1 2.2 . High RDW level in the blood. MrLabTest. Last update: 12/01/2021
  3. 3.0 3.1 Teresa Scordino (2016-12-02). Giant platelets. American Society of Hematology.
  4. Glassy, Eric (1998). Color atlas of hematology : an illustrated field guide based on proficiency testing . Northfield, Ill: College of American Patholgists. ISBN 978-0-930304-66-9. OCLC 40976106. 
  5. . prolymphocytes in PLL. American Society of Hematology (2013-07-16).
  6. Fredrick L. Kiechle. Q & A. CAP Today. June 2010

Image sources


Platelet aggregation study

Author: Mikael Häggström [note 1]

On for example optical densitometry, a first and second wave of platelet aggregation is seen, in this case for an ADP-initiated aggregation.[1]

In a platelet aggregation study, the aggregation process is started by different agonists (ADP, epinephrine etc.) and the aggregation pattern can usually conform into any of the following patterns:

Platelet aggregation function by disorders and agonists
ADP Epinephrine Collagen Ristocetin
P2Y receptor defect[2] (including Clopidogrel) Decreased Normal Normal Normal
Adrenergic receptor defect[2] Normal Decreased Normal Normal
Collagen receptor defect[2] Normal Normal Decreased or absent Normal
  • Von Willebrand disease[3]
  • Bernard–Soulier syndrome[2]
Normal Normal Normal Decreased or absent
  • Glanzmann's thrombasthenia[2]
  • Afibrinogenemia
Decreased Decreased Decreased Normal or decreased
Storage pool deficiency[3] Absent second wave Partial
Aspirin or aspirin-like disorder Absent second wave Absent Normal

Further reading

Notes

  1. For a full list of contributors, see article history. Creators of images are attributed at the image description pages, seen by clicking on the images. See Patholines:Authorship for details.

Main page

References

  1. Jiang, L.; Xu, C.; Yu, S.; Liu, P.; Luo, D.; Zhou, Q.; Gao, C.; Hu, H. (2013). "A critical role of thrombin/PAR-1 in ADP-induced platelet secretion and the second wave of aggregation ". Journal of Thrombosis and Haemostasis 11 (5): 930–940. doi:10.1111/jth.12168. ISSN 15387933. PMID 23406164. 
  2. 2.0 2.1 2.2 2.3 2.4 Borhany, Munira; Pahore, Zaen; ul Qadr, Zeeshan; Rehan, Muhammad; Naz, Arshi; Khan, Asif; Ansari, Saqib; Farzana, Tasneem; et al. (2010). "Bleeding disorders in the tribe: result of consanguineous in breeding ". Orphanet Journal of Rare Diseases 5 (1). doi:10.1186/1750-1172-5-23. ISSN 1750-1172. 
  3. 3.0 3.1 . Why Perform Platelet Aggregation?. Helena Biosciences. 2015

Image sources


Bone marrow

Author: Mikael Häggström [note 1]

Bone marrow aspirate showing normal "trilineage hematopoiesis":
- Myelomonocytic cells: an eosinophil myelocyte marked
- Erythroid cells: an orthochromatic erythroblast marked
- Megakaryocytic cells.

Autopsy

Using pliers or similar tool, squeeze some bone marrow from a rib.

Microscopic evaluation

  • Confirm trilineage hematopoiesis (see image).
  • Look for apparent cellular atypia or decrease of cellular diversity.

Report

Example in a normal case:

Bone marrow from rib (or other location if applicable): Trilineage hematopoiesis. There is no evidence of malignancy.

Bone marrow biopsy

Gross processing

  • Ensure that the biopsy is properly fixed (generally at least 2 hours).
  • Measure length and diameter of each fragment.
  • If the biopsy is received in a zinc-containing fixative, rinse it for about 2 minutes, such as placing the cassette in a container under running water (and block any sink enough so that the cassette won't float away).
Histopathology of bone marrow with insufficient decalcification, wherein the bony trabeculae may get pushed by the microtome blade and plow away the cells of interest as displayed. To compensate, thicker sections may be taken, also making evaluation harder.
  • Put in decalcifying solution, preferably using a type that is optimal for performing subsequent immunohistochemistry. Generally decalcify the specimen for about 15 minutes initially, and palpate the specimen to check whether it is still firm. If it is, decalcify another 5-10 minutes and check again. Rather have it a little bit too soft than a little bit too firm. Be careful to follow grossing guidelines for bone marrows, since even a small aberration in the processing is likely to be noticed as creating artifacts.
Gross report example
The specimen is received in AZF solution and consists of __ core needle bone marrow biopsy fragment(s) measuring __ cm in length and 0.2 cm in diameter. The specimen is entirely submitted for microscopic examination in one cassette following decalcification.

Microscopic evaluation

Evaluate the following:

Bone marrow aspirate

Find a location where bone marrow cells can be seen individually, preferably with minimal peripheral blood among them. Sometimes it is between trabeculae and sometimes it is in the surrounding area. Perform a differential count by counting 500 cells[1] and dividing the numbers by 5 to get their percentages. Don't count cells that don't have a cytoplasm or nucleus. If you receive several slides, count about the same number of cells from each slide, but don't count cells from slides where the cell types are more indistinct.

  • Myeloid/erythroid ratio, usually defined as the ratio between the number of neutrophil granulocytes and precursors versus the number of erythroblasts. Depending on source, the lower limit of the normal range for this ratio bone marrow aspirate smears in adults is 1 to 2, and the upper limit is 5 to 8.[2] In histologic sections, the normal range is 1.5 – 3.0.[2] Some hematologists also include eosinophils, basophils and monocytes, as well as their precursors, in the myeloid number, but this has only a minor effect on the M:E ratio in normal individuals.[2] Still, if you are to calculate the value for a senior, check their preference first.

In a bone marrow count, nucleated red blood cells count into the total percentage of cells (but does not count into percentage of white blood cells in peripheral blood).

Bone marrow biopsy, H&E stain
  • Adequacy: There should preferably be 5 intertrabecular spaces.
  • Cellularity: This is a rather rough estimation of the area of hematopoietic cells divided by the area of all intertrabecular matter, which otherwise mainly consists of fat cells and a small interstitial space. Areas of bone, hemorrhage or artifacts are not counted. In patients 20-80 years, the percentage should be about 100 minus age, such as 40% in a 60 year old patient.
  • Thrombopoiesis: There are normally about 3 megakaryocytes per 40x field.
  • Look for any granuloma or cancer metastasis
Bone marrow biopsy, other stains

Generally including:

Ring sideroblast, defined by five or more perinuclear iron granules that encompass at least 1/3 of the nuclear circumference.[3]
  • Iron stain: Look at approximately 100 cells and semi-quantify the presence of any ring sideroblasts.
  • Reticulin to grade the amount of fibrosis.
  • CD3 and CD20 to highlight T and B cells, respectively.

Microscopic report

Example template:

On this resource, the following formatting is used for comprehensiveness:

  • Minimal depth
  • (Moderate depth)
  • ((Comprehensive))
Other legend

<< Decision needed between alternatives separated by / signs >>
{{Common findings / In case of findings}}
[[Comments]]
Link to another page

SOURCE:

Bone marrow aspirate and biopsy

CLINICAL INFORMATION:

{{Lymphocytosis / Pancytopenia / Anemia etc }}

DIAGNOSIS:
PERIPHERAL BLOOD:

{{Lymphocytosis / Pancytopenia / Anemia etc }}

BONE MARROW, {{POSTERIOR ILIAC CREST,}} ASPIRATE AND BIOPSY:

{{Chronic/Acute lymphocytic/myeloid leukemia.}}
See description and comment.

GROSS DESCRIPTION:

[[As per gross report above.]]

MICROSCOPIC DESCRIPTION:

PERIPHERAL SMEAR:

MORPHOLOGY:

  • Red Blood Cells: No significant abnormality.
  • White Blood Cells: No significant abnormality.
  • Platelets: No significant abnormality.

BONE MARROW ASPIRATE:

Left / RIGHT / UNDESIGNATED SITE

CELLULARITY ESTIMATE: Adequate. / Hypocellular and hemodilute. / Hypercellular. / Too few cells for morphologic evaluation.

MARROW DIFFERENTIAL

  • Erythroblasts: __%
  • Blasts: __%
  • Neutrophils/precursors: __%
  • Eosinophils: __%
  • Basophils: __%
  • Lymphocytes: __%
  • Monocytes: __%
  • Plasma Cells: __%
  • M:E ratio: __:__

ERYTHROPOIESIS: Maturing.

GRANULOPOIESIS: Maturing.

MEGAKARYOCYTES: Present.

LYMPHOCYTES: Mature.

PLASMA CELLS: Rare without atypia.

BONE MARROW TOUCH PREPARATION: _

BONE MARROW BIOPSY:

Left / RIGHT / UNDESIGNATED SITE

SPECIMEN ADEQUACY:

Satisfactory. / Limited. / Unsatisfactory.

CELLULARITY:_

_%: Normocellular / Hypercellular / Hypocellular for age. / Variable ( _ %- _ %, overall _ %) / Cannot assess cellularity due to small biopsy. {{With aspirative artifact.}}

Decalcified bone marrow biopsy demonstrates ____________________

BM Erythropoiesis _

BM Cellularity _

BM Megakaryocytes _

BM Myelopoiesis _

BONE MARROW IRON STAIN: Storage iron _/4 on aspirate smear with / without ring sideroblasts.

SPECIAL STAINS:

Reticulin Stain: ___ reticulin fibrosis (_+/3+)

FLOW CYTOMETRY:

No immunophenotypic evidence for abnormal myeloid maturation, an increase in blasts, or a lymphoproliferative disorder (see attached report).

COMMENT:

{{Cytogenetic study and next generation sequencing panel for myeloid neoplasm are pending.}}
All stains have functional controls.


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  1. Abdulrahman AA, Patel KH, Yang T, Koch DD, Sivers SM, Smith GH (2018). "Is a 500-Cell Count Necessary for Bone Marrow Differentials?: A Proposed Analytical Method for Validating a Lower Cutoff. ". Am J Clin Pathol 150 (1): 84-91. doi:10.1093/ajcp/aqy034. PMID 29757362. Archived from the original. . 
  2. 2.0 2.1 2.2 BJ. Bain (2017-02-19). Chapter 5: Pathology of the marrow. Basicmedical Key.
  3. Salama M, Teruya-Feldstein J, Kremyankaya M. Atlas of Diagnostic Hematology. Philadelphia, PA: Elsevier; 2021.