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immunophenotypic diagnosis of CLL
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file:PC.jpg|link={{filepath:PC.jpg}}
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'''Background'''


CLL is a neoplasm of B-lymphocytes whose distinctive biology often allows confident diagnosis:
:First, the cells are "anergic" i.e. they have down-regulated their B-cell receptor (BCR) and its associated molecules - so immunoglobulin light chains, CD19, CD20 and particularly CD79b are all weakly expresssed.
:Second, the cells generally express CD5 and CD23 (which are normally expressed on only a small proporton of normal B-cells) which gives them a relatively unique immunophenotypic profile


Consequently '''tyical'' CLL can be diagnosed with some confidence using flow cytometry. However, no single marker is abolutely typical, and atypical cases are regularly encoutered. These atypical cases can cause diagnosic difficulty.
<span style="color: navy>'''Background to flow cytometric diagnosis'''</span>


file CD45CLL.jpg left 250px 
Differentiation to plasma cells involves (in mast cases) the loss of typical pan B-cell markers and surface light chains. The expression of CD45 also differs both within and between cases making standard gating strategies difficult to apply. Cells are most often identified through their co-expression of CD138 with a relatively high expression of CD38, together with the absence of typical B-cell features.


''This image shows a typical CD45 ssc plot of CLL. Note that although the CLL cells (shown in blue) generally lie close to the position of normal lymphocytes, consistent with their anergic natures they will generally have lower expression of CD45 than normal typical T cells (shown in red) or NK cells (shown in green). They also have wider range of CD45 expression causing them to have a "sausage like" shape. This is a fairly typical appearance of CLL though not in itself diagnostic.''


'''Diagnosis of CLL''


* CD19 should be detected but intensity is generally low in CLL
<div style="width: 95%; overflow: auto; border: 1px solid navy; font-size:100%">
* CD5 should be expressed at moderate intensity in CLL (note also on expressed on mcl and occasionally on other lpd) 
{| class="wikitable" style="color:black; background-color:#ffffff;" cellpadding="5"
* CD23 has expected expression in CLL often at low intensity, generally absent in mcl but variably present on other lpd
!colspan="4" style = "font-size:110%; color: navy; text-align:center; background:#DEE0DE;"|'''Immunophenotype of plasma cells'''
* CD79b similar to CD19 this marker is expected to be expressed but intensity is generally low in CLL
|-
* immunoglobulin light chains sig expect either light chain restriction with low intensity or undetectable
!colspan="4" style = "font-size:90%; background:#DAE8F2"|'''CD45''' expression
* fmc7 a marker of b-cell activation that should have low or absent expression, high expression is particularly seen in pll and hairy cell-like disorders
|-
* CD200 expressed on most CLL cases as well as other b-lpd but is most frequently absent in mcl  optional markers extended panel for atypical cases 
|colspan="4" style = "font-size:90%; color:black;" |There is no consistent pattern of expression of CD45 although it is most often weak, but may be variable and is occasionally strong.
* CD20 a pan b-cell marker with expected expression in CLL but intensity is generally low
|-
* CD43 a pan b-cell marker with expected expression in CLL; frequently but not always absent in mcl
!colspan="4" style = "font-size:90%; background:#DAE8F2"|Major markers useful in plasma cell identification
|-
|colspan="1" style = "font-size:80%; width: 4%; |'''Marker'''
|colspan="1" style = "font-size:80%; width: 8%; |'''Freq'''
|colspan="1" style = "font-size:80%; width: 8%; |'''Level'''
|colspan="1" style = "font-size:80%; width: 80%; |'''Comment'''
|-
|colspan="1" style = "font-size:90%; color:black;" |CD38
|colspan="1" style = "background:#004466;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|The relatively strong expression of CD38 is typical of plasma cells
|-
|colspan="1" style = "font-size:90%;" |CD138
|colspan="1" style = "background:#004466;"|
|colspan="1" style = "font-size:80%;"|var
|colspan="1" style = "font-size:84%;"|Expression is expected although the intensity can vary
|-
|colspan="1" style = "width: 4%; font-size:90%; color:black;" |κ/λ
|colspan="1" style = "width: 8%; background:#E6FAFF;"|
|colspan="1" style = "font-size:80%; width: 8%; |
|colspan="1" style = "font-size:80%;"|Surface light chain expression is lost (internal will be detected in permeabilised cells)
|-
|colspan="1" style = "font-size:90%; color:black;" |CD19
|colspan="1" style = "background:#66e0ff;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|Expression of CD19 may persist in some cases  
|-
!colspan="4" style = "font-size:90%; background:#DAE8F2"|Other relevant markers  
|-
|colspan="1" style = "font-size:90%; color:black;" |CD5
|colspan="1" style = "background:#E6FAFF;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:80%;"|Not detected on plasma cells
|-
|colspan="1" style = "font-size:90%;" |CD10
|colspan="1" style = "background:#E6FAFF;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|
|-
|colspan="1" style = "font-size:90%;" |CD11C
|colspan="1" style = "background:#E6FAFF;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|
|-
|colspan="1" style = "font-size:90%;" |CD20
|colspan="1" style = "background:#66e0ff;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|Expression retained in some cases
|-
|colspan="1" style = "font-size:90%; color:black;" |CD23
|colspan="1" style = "background:#E6FAFF;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|
|-
|colspan="1" style = "font-size:90%; color:black;" |CD25
|colspan="1" style = "background:#E6FAFF;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|
|-
|colspan="1" style = "font-size:90%;" |CD43
|colspan="1" style = "background:#006699;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|Generally not expressed by LPL
|-
|colspan="1" style = "font-size:90%; color:black;" |CD79b
|colspan="1" style = "background:#66e0ff;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|Expressed in some cases
|-
|colspan="1" style = "font-size:90%;" |CD103
|colspan="1" style = "background:#E6FAFF;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|
|-
|colspan="1" style = "font-size:90%; color:black;" |CD200
|colspan="1" style = "background:#006699;"|
|colspan="1" style = "font-size:80%;"|-
|colspan="1" style = "font-size:84%;"|Detected frequently|-|}
|-
|colspan="1" style = "font-size:90%; color:black;" |FMC7
|colspan="1" style = "background:#E6FAFF;"|
|colspan="1" style = "font-size:80%;"|
|colspan="1" style = "font-size:80%;"|Not expressed
|-
|}
</div>
 
 
 
<span style="color:navy;font-size:80% ">'''Key to table: '''</span>
 
<span style="font-size:80% ">Key to colour code for expression frequency [[Key_to_marker_expression_table|Click for link]]</span>
 
<span style="font-size:80% ">Key to expression strength code and use [[Expression strength key|Click for link]]</span>
 
 
 
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Latest revision as of 18:09, 14 July 2023

  • PC.jpg


Background to flow cytometric diagnosis

Differentiation to plasma cells involves (in mast cases) the loss of typical pan B-cell markers and surface light chains. The expression of CD45 also differs both within and between cases making standard gating strategies difficult to apply. Cells are most often identified through their co-expression of CD138 with a relatively high expression of CD38, together with the absence of typical B-cell features.


Immunophenotype of plasma cells
CD45 expression
There is no consistent pattern of expression of CD45 although it is most often weak, but may be variable and is occasionally strong.
Major markers useful in plasma cell identification
Marker Freq Level Comment
CD38 - The relatively strong expression of CD38 is typical of plasma cells
CD138 var Expression is expected although the intensity can vary
κ/λ Surface light chain expression is lost (internal will be detected in permeabilised cells)
CD19 - Expression of CD19 may persist in some cases
Other relevant markers
CD5 - Not detected on plasma cells
CD10 -
CD11C -
CD20 - Expression retained in some cases
CD23 -
CD25 -
CD43 - Generally not expressed by LPL
CD79b - Expressed in some cases
CD103 -
CD200 - Detected frequently|-|}
FMC7 Not expressed


Key to table:

Key to colour code for expression frequency Click for link

Key to expression strength code and use Click for link


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