Katarina Junevik
Katarina Junevik
Katarina Junevik
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Lymfomdiagnostik med<br />
flödescytometri<br />
Diagnostikforum<br />
Jönköping 2012-19-27<br />
<strong>Katarina</strong> <strong>Junevik</strong>, leg Biomedicinsk analytiker<br />
Laboratoriet för klinisk kemi,<br />
Sahlgrenska Universitetssjukhus Göteborg
Flödescytometri – singelceller i<br />
ett flöde som passerar en laser<br />
eller två<br />
eller tre<br />
eller sju........<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Vad kan flödescytometern ge oss<br />
för information om cellen<br />
SSC<br />
+ fluorescens<br />
FSC<br />
Dess relativa storlek - Forward Scatter FSC<br />
Dess granulering eller inre komplextivitet - Side Scatter<br />
SSC<br />
Dess relativa fluorescens intensitet<br />
<strong>Katarina</strong> <strong>Junevik</strong>
FSC/SSC<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Cellulära antigen<br />
Membran-receptor<br />
– Receptorer<br />
– Adhesionsmolekyler<br />
– Enzymer<br />
Nukleära antigen<br />
Cytoplasmatiska<br />
antigen<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Permeabilisering<br />
För att kunna märka in intracellulära<br />
antigen/cytokiner permeabiliseras cellen.<br />
Antikropp<br />
ic antigen<br />
Permeabilisering<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Direktkonjungering<br />
Fluorokrom<br />
Monoklonal antikropp<br />
med konjungerad<br />
fluorokrom<br />
Antikroppsdel<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Indirekt inmärkning<br />
Sekundär ak<br />
råtta anti-mus<br />
Primär ak<br />
mus anti-human<br />
Primär antikropp:<br />
okonjungerad<br />
Sekundär antikropp:<br />
direktkonjungerad<br />
Human cell<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Cluster of Differentiation<br />
CD<br />
organisation<br />
CD<br />
organisation<br />
Ak<br />
lab<br />
Cell<br />
lab<br />
CD<br />
workshop<br />
CD nr<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Fluorescens<br />
FITC<br />
FITC<br />
FITC<br />
FITC<br />
FITC<br />
FITC<br />
FITC<br />
FITC<br />
FITC<br />
FITC<br />
Antal<br />
Fluorescens intensitet<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Material<br />
Prov<br />
– Blod<br />
– Benmärg<br />
– Likvor<br />
– Ascites<br />
– Pleura<br />
– Biopsi (lymfkörtel)
www.euroflow.org<br />
EuroFlow Antibody<br />
Panels<br />
Uniform Coding of<br />
Panels<br />
EuroFlow Marker<br />
List<br />
Diagnostic Diagram
B-CLPD (B-cell lympho proliferative disease)<br />
V-450 V-<br />
CD20 + CD4<br />
2µl + 2µl<br />
CD45<br />
2µl<br />
50<br />
0<br />
FITC PE PerCP -<br />
Cy5.5<br />
CD8+lambda CD56+kappa<br />
5µl<br />
CD5<br />
5µl<br />
PC7 APC APC-<br />
H7<br />
CD19+TCRγδ<br />
5µl + 2µl<br />
CD3<br />
2µl<br />
CD38<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD23<br />
5µl<br />
CD10<br />
5µl<br />
CD79b<br />
2µl<br />
CD19<br />
5µl<br />
CD200<br />
2µl<br />
CD43<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD31<br />
5µl<br />
LAIR1<br />
5µl<br />
CD11c<br />
2µl<br />
CD19<br />
5µl<br />
IgM<br />
5µl<br />
CD81<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD103<br />
5µl<br />
CD95<br />
5µl<br />
CD22<br />
2µl<br />
CD19<br />
5µl<br />
CXCR5<br />
5µl<br />
CD49d<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD62L<br />
5µl<br />
CD39<br />
5µl<br />
HLA-DR<br />
5µl<br />
CD19<br />
5µl<br />
CD27<br />
2µl<br />
CD25<br />
5µl<br />
<strong>Katarina</strong> <strong>Junevik</strong>
RÖR 1
B-CLPD (B-cell lympho proliferative disease)<br />
V-450 V-<br />
CD20 + CD4<br />
2µl + 2µl<br />
CD45<br />
2µl<br />
50<br />
0<br />
FITC PE PerCP -<br />
Cy5.5<br />
CD8+lambda CD56+kappa<br />
5µl<br />
CD5<br />
5µl<br />
PC7 APC APC-<br />
H7<br />
CD19+TCRγδ<br />
5µl + 2µl<br />
CD3<br />
2µl<br />
CD38<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD23<br />
5µl<br />
CD10<br />
5µl<br />
CD79b<br />
2µl<br />
CD19<br />
5µl<br />
CD200<br />
2µl<br />
CD43<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD31<br />
5µl<br />
LAIR1<br />
5µl<br />
CD11c<br />
2µl<br />
CD19<br />
5µl<br />
IgM<br />
5µl<br />
CD81<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD103<br />
5µl<br />
CD95<br />
5µl<br />
CD22<br />
2µl<br />
CD19<br />
5µl<br />
CXCR5<br />
5µl<br />
CD49d<br />
2µl<br />
CD20<br />
2µl<br />
CD45<br />
2µl<br />
CD62L<br />
5µl<br />
CD39<br />
5µl<br />
HLA-DR<br />
5µl<br />
CD19<br />
5µl<br />
CD27<br />
2µl<br />
CD25<br />
5µl<br />
<strong>Katarina</strong> <strong>Junevik</strong>
RÖR 2
Lymfom<br />
Normalblod<br />
KLL<br />
Mantelcells<br />
lymfom<br />
Follikulärt<br />
lymfom<br />
CD20<br />
CD19<br />
lambda<br />
kappa<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Normalblod<br />
Lymfom forts.<br />
KLL<br />
Mantelcells<br />
lymfom<br />
Follikulärt<br />
lymfom<br />
CD5<br />
CD20<br />
CD20<br />
CD23<br />
CD20<br />
CD10<br />
<strong>Katarina</strong> <strong>Junevik</strong>
SST (small sample tube)<br />
V 450 V 500 FITC PE PerCP<br />
-Cy5.5<br />
PC7 APC APC-<br />
H7<br />
CD20 CD45 CD8 + lambda CD56 + kappa CD4 CD19 CD3+CD14 CD38<br />
<strong>Katarina</strong> <strong>Junevik</strong>
Tack för uppmärksamheten ☺
T-CLPD<br />
V450 V500 FITC PE PerCP-<br />
Cy5.5<br />
PE-Cy7 APC APC-H7<br />
CD4 CD45 CD7 CD26 CD3 CD2 CD28 CD8<br />
CD4 CD45 CD27 CCR7 CD3 CD45RO CD45RA CD8<br />
CD4 CD45 CD5 CD25 CD3 HLADR ic TCL1 CD8<br />
CD4 CD45 CD57 CD30 CD3 CD11c CD8<br />
CD4 CD45 ic Perforin ic Granzyme CD3 CD16 CD94 CD8<br />
CD4 CD45 CD279 CD3 CD8
Myelom<br />
V 450 V 500 FITC PE PerCP-<br />
Cy5.5<br />
PC7 APC APC-H7<br />
CD138 CD45 CD38 CD56 BTA2micro CD19 Kappa<br />
ic<br />
Lambda<br />
ic<br />
CD138 CD45 CD38 CD28 CD27 CD19 CD117 CD81<br />
NKcellspanel<br />
V450 V500 FITC PE PerCP-<br />
Cy5.5<br />
CD2<br />
Pacific Blue<br />
5µL<br />
CD16<br />
2µL<br />
HLA-DR<br />
2µL<br />
CD45<br />
2µL<br />
CD45<br />
2µL<br />
CD45<br />
2µL<br />
CD7<br />
5µL<br />
CD57<br />
5µL<br />
ic<br />
Perforin<br />
5µL<br />
CD26<br />
5µL<br />
CD25<br />
5µL<br />
ic<br />
Granzyme<br />
2µL<br />
CD3<br />
5µL<br />
CD3<br />
5µL<br />
CD3<br />
5µL<br />
PC7 APC APC-H7<br />
CD56<br />
2µL<br />
CD56<br />
2µL<br />
CD56<br />
2µL<br />
CD5<br />
2µL<br />
CD11c<br />
BD<br />
2µL<br />
CD94<br />
5µL<br />
CD19<br />
5µL<br />
CD19<br />
5µL<br />
CD19<br />
5µL
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