Immunohistochemistry: Difference between revisions

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When [[learning pathology]], the percentages by which immunohistochemistry results are positive or negative for various diseases are generally easily looked up when needed, so what a pathologist needs to learn is mainly '''how to select''' the optimal immunohistochemistry panels in the first place for various presentations where the diagnosis is unknown.

==Immunohistochemistry ordering==

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*Consider additional stains from the '''"Comprehensive panel"''' displayed below the suggested one.

*Switch the '''"Sensitivity"''' setting (seen at top) from 1 (which means that diffuse, focal as well as not specified staining count as positive) to 3 (which means that only diffuse staining counts as positive whereas both focal and absent staining count as negative, and references without any specified staining pattern are omitted from the analysis). This has less data to support the suggested stains (since many references do not specify whether positivity was diffuse or focal), but can sometimes state a better distinction between conditions. When including a stain based on its distinguishing features on a sensitivity setting of 3, you need to keep the practice of classifying only diffuse staining counts as positive, and focal to absent staining as negative.

===Test question: ImmunoQuery for a squamous cell carcinoma===

[[File:Histopathology of squamous-cell carcinoma of the lung.jpg|thumb|230px|Squamous cell carcinoma, with large cells with abundant eosinophilic cytoplasm and large, often vesicular, nuclei.]]

The attending gives you a lung biopsy case to preview. You are first uncertain about the type of tumor, so you ask a fellow resident, who finds a diagnostic area of the tumor and tells you that this is a typical squamous cell carcinoma. You also look through the patient's history, and find that the patient has had a squamous cell carcinoma of the anus in the past, and you now want to find out whether the tumor originated in the lung, or if it is a metastasis from the anus, or possibly the skin. You therefore do an ImmunoQuery lookup, with the following results:

;Suggested Panel

''Insufficient antibodies for a satisfactory panel to differentiate Lung squamous cell carcinoma and Anus squamous cell carcinoma''

{|class=wikitable

| Antibodies

! Lung SCC !! Anus SCC !! Skin SCC

|-

! EpCAM

| 81% Positive Membrane, Cytoplasm || 75% Positive Membrane, Cytoplasm || 0% Positive Membrane, Cytoplasm

|-

! GATA3

| 5% Positive Nucleus || 20% Positive Nucleus || 84% Positive Nucleus

|-

! p16

| 17% Positive Cytoplasm, Nucleus || 87$ Positive Cytoplasm, Nucleus || 45% Positive Cytoplasm, Nucleus

|}

You go talk with the attending, who agrees that EpCAM, GATA3 and p16 should be in the panel, but just as ImmunoQuery also tells, the attending thinks that the panel is not satisfactory to differentiate lung SCC from anus SCC, and wants you to add one more stain to improve the panel. You go back to ImmunoQuery and increase the Sensitivity from 1 to 3, and get the following results:

;Suggested panel

{|class=wikitable

| Antibodies

! Lung SCC !! Anus SCC !! Skin SCC

|-

! EpCAM

| 74% Positive Membrane, Cytoplasm || 50% Positive Membrane, Cytoplasm || 0% Positive Membrane, Cytoplasm

|-

! DLK

| 28% Positive Membrane, Cytoplasm || N/A Membrane, Cytoplasm || 100% Positive Membrane, Cytoplasm

|}

You also perform a repeated search by only entering Lung and Anus SCC, and you get the following results:

;Suggested panel

{|class=wikitable

| Antibodies

! Lung SCC !! Anus SCC

|-

! p16

| 17% Positive Cytoplasm, Nucleus || 87% Positive Cytoplasm, Nucleus

|}

;Comprehensive panel

Top result:

{|class=wikitable

| Antibodies

! Lung SCC !! Anus SCC

|-

| '''GRPR''' Cytoplasm || 46% || 91%

|}

You go talk with a technician at the histology lab, and your hospital offers all the stains in the alternatives, at similar costs, so you don't have to think about expenses and logistics of sending the case out to external labs.

What is the best alternative?

*Choose Cyklin-D1, and favor a lung primary if the stain has even just focal positivity.

*Choose Cyklin-D1, and favor a lung primary if the stain is diffuse rather than focal or no reactivity. (correct)

- Choose EGFR

==Immunohistochemistry evaluation==

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*Paying for a subscription to '''ImmunoQuery'''<ref group=notes name=ImmunoQueryCOI/>, where you can enter immunohistochemistry results and generate a list of most likely conditions with that profile.

Preferably, immunohistochemistry results will be very specific or sensitive for a suspected condition, thereby confirming it if positive, or excluding it if negative, respectively. Even when that is not the case, immunohistochemistry can at least alter the likelihoods of different differential diagnoses. Pathology practice is too uncertain to perform calculations of exact percentages of likelihoods of differential diagnoses, but to demonstrate the general principle of how immunohistochemistry results are calculated, the following formula can be used:

Preferably, immunohistochemistry results will be very specific or sensitive for a suspected condition, thereby confirming it if positive, or excluding it if negative, respectively. Even when that is not the case, immunohistochemistry can at least '''alter the likelihoods''' of different differential diagnoses. In practice, clinicians or pathologists do not state exact or even approximate numbers of likelihoods of differential diagnoses ({{further|Reporting}}<includeonly>see the [[Reporting]] chapter for phrasing uncertainty</includeonly>), since reality is too complex for that, but the following mathematical should still be somewhat followed in order to interpret immunohistochemistry results optimally:

In practice, the most feasible

Pathology practice is too uncertain to perform calculations of exact percentages of likelihoods of differential diagnoses, but to demonstrate the general principle of how immunohistochemistry results are calculated, the following formula can be used:

*Gross likelihood of a disease/condition = (Pre-test probability) x (Probability that the condition shows the immunohistochemistry results at hand).

The pre-test probability is a product of for example the incidence of the condition in the patient's epidemiologic type such as age and sex, as well as the probability that the condition would have caused the clinical course, including signs and symptoms, as well as the microscopic impression. For example, if you want to differentiate a pleomorphic liposarcoma from a pleomorphic rhabdomyosarcoma in soft tissue, you may find in ImmunoQuery that the following stains are most efficient in distinguishing the two, with the following percentages of being positive:

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 |author2=
 }}
 When [[learning pathology]], the percentages by which immunohistochemistry results are positive or negative for various diseases are generally easily looked up when needed, so what a pathologist needs to learn is mainly '''how to select''' the optimal immunohistochemistry panels in the first place for various presentations where the diagnosis is unknown.
 ==Immunohistochemistry ordering==
@@ Line 15: / Line 15: @@
 *Consider additional stains from the '''"Comprehensive panel"''' displayed below the suggested one.
 *Switch the '''"Sensitivity"''' setting (seen at top) from 1 (which means that diffuse, focal as well as not specified staining count as positive) to 3 (which means that only diffuse staining counts as positive whereas both focal and absent staining count as negative, and references without any specified staining pattern are omitted from the analysis). This has less data to support the suggested stains (since many references do not specify whether positivity was diffuse or focal), but can sometimes state a better distinction between conditions. When including a stain based on its distinguishing features on a sensitivity setting of 3, you need to keep the practice of classifying only diffuse staining counts as positive, and focal to absent staining as negative.
+===Test question: ImmunoQuery for a squamous cell carcinoma===
+[[File:Histopathology of squamous-cell carcinoma of the lung.jpg|thumb|230px|Squamous cell carcinoma, with large cells with abundant eosinophilic cytoplasm and large, often vesicular, nuclei.]]
+The attending gives you a lung biopsy case to preview. You are first uncertain about the type of tumor, so you ask a fellow resident, who finds a diagnostic area of the tumor and tells you that this is a typical squamous cell carcinoma. You also look through the patient's history, and find that the patient has had a squamous cell carcinoma of the anus in the past, and you now want to find out whether the tumor originated in the lung, or if it is a metastasis from the anus, or possibly the skin. You therefore do an ImmunoQuery lookup, with the following results:
+;Suggested Panel
+''Insufficient antibodies for a satisfactory panel to differentiate Lung squamous cell carcinoma and Anus squamous cell carcinoma''
+{|class=wikitable
+| Antibodies
+! Lung SCC !! Anus SCC !! Skin SCC
+|-
+! EpCAM
+| 81% Positive<br>Membrane, Cytoplasm || 75% Positive<br>Membrane, Cytoplasm || 0% Positive<br>Membrane, Cytoplasm
+|-
+! GATA3
+| 5% Positive<br>Nucleus || 20% Positive<br>Nucleus || 84% Positive<br>Nucleus
+|-
+! p16
+| 17% Positive<br>Cytoplasm, Nucleus || 87$ Positive<br>Cytoplasm, Nucleus || 45% Positive<br>Cytoplasm, Nucleus
+|}
+You go talk with the attending, who agrees that EpCAM, GATA3 and p16 should be in the panel, but just as ImmunoQuery also tells, the attending thinks that the panel is not satisfactory to differentiate lung SCC from anus SCC, and wants you to add one more stain to improve the panel. You go back to ImmunoQuery and increase the Sensitivity from 1 to 3, and get the following results:
+;Suggested panel
+{|class=wikitable
+| Antibodies
+! Lung SCC !! Anus SCC !! Skin SCC
+|-
+! EpCAM
+| 74% Positive<br>Membrane, Cytoplasm || 50% Positive<br>Membrane, Cytoplasm || 0% Positive<br>Membrane, Cytoplasm
+|-
+! DLK
+| 28% Positive<br>Membrane, Cytoplasm || N/A<br>Membrane, Cytoplasm || 100% Positive<br>Membrane, Cytoplasm
+|}
+You also perform a repeated search by only entering Lung and Anus SCC, and you get the following results:
+;Suggested panel
+{|class=wikitable
+| Antibodies
+! Lung SCC !! Anus SCC
+|-
+! p16
+| 17% Positive<br>Cytoplasm, Nucleus || 87% Positive<br>Cytoplasm, Nucleus
+|}
+;Comprehensive panel
+Top results:
+{|class=wikitable
+| Antibodies
+! Lung SCC !! Anus SCC
+|-
+| '''p16'''<br>Cytoplasm, Nucleus || 17% || 87%
+|-
+| '''GRPR'''<br>Cytoplasm || 56% || 100%
+|}
+You switch sensitivity from 1 to 3 for this result as well, showing:
+;Suggested panel
+''No antibodies to differentiate Lung squamous cell carcinoma and Anus squamous cell carcinoma''
+;Comprehensive panel
+Top result:
+{|class=wikitable
+| Antibodies
+! Lung SCC !! Anus SCC
+|-
+| '''GRPR'''<br>Cytoplasm || 46% || 91%
+|}
+You go talk with a technician at the histology lab, and your hospital offers all the stains in the alternatives, at similar costs, so you don't have to think about expenses and logistics of sending the case out to external labs.
+What is the best alternative?
+*Choose Cyklin-D1, and favor a lung primary if the stain has even just focal positivity.
+*Choose Cyklin-D1, and favor a lung primary if the stain is diffuse rather than focal or no reactivity.  (correct)
+- Choose EGFR
 ==Immunohistochemistry evaluation==
@@ Line 21: / Line 92: @@
 *Paying for a subscription to '''ImmunoQuery'''<ref group=notes name=ImmunoQueryCOI/>, where you can enter immunohistochemistry results and generate a list of most likely conditions with that profile.
-Preferably, immunohistochemistry results will be very specific or sensitive for a suspected condition, thereby confirming it if positive, or excluding it if negative, respectively. Even when that is not the case, immunohistochemistry can at least alter the likelihoods of different differential diagnoses. Pathology practice is too uncertain to perform calculations of exact percentages of likelihoods of differential diagnoses, but to demonstrate the general principle of how immunohistochemistry results are calculated, the following formula can be used:
+Preferably, immunohistochemistry results will be very specific or sensitive for a suspected condition, thereby confirming it if positive, or excluding it if negative, respectively. Even when that is not the case, immunohistochemistry can at least '''alter the likelihoods''' of different differential diagnoses. In practice, clinicians or pathologists do not state exact or even approximate numbers of likelihoods of differential diagnoses ({{further|Reporting}}<includeonly>see the [[Reporting]] chapter for phrasing uncertainty</includeonly>), since reality is too complex for that, but the following mathematical should still be somewhat followed in order to interpret immunohistochemistry results optimally:
+In practice, the most feasible
+Pathology practice is too uncertain to perform calculations of exact percentages of likelihoods of differential diagnoses, but to demonstrate the general principle of how immunohistochemistry results are calculated, the following formula can be used:
 *Gross likelihood of a disease/condition = (Pre-test probability) x (Probability that the condition shows the immunohistochemistry results at hand).
 The pre-test probability is a product of for example the incidence of the condition in the patient's epidemiologic type such as age and sex, as well as the probability that the condition would have caused the clinical course, including signs and symptoms, as well as the microscopic impression. For example, if you want to differentiate a pleomorphic liposarcoma from a pleomorphic rhabdomyosarcoma in soft tissue, you may find in ImmunoQuery that the following stains are most efficient in distinguishing the two, with the following percentages of being positive:

Immunohistochemistry: Difference between revisions

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