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Selective stain of mutated CALR protein in megacaryocytes of early primary myelofibrosis (anti-CALRmut clone CAL2 1:50, Dako Omnis, 400x).
Selective stain of mutated CALR protein in megacaryocytes of essential thrombocythaemia (anti-CALRmut clone CAL2 1:50, Dako Omnis, 200x).
CAL2 - a monoclonal antibody for detection of all types of CALR mutations in MPNs (CALRmut-PMF, Dako Omnis, CAL2, 1-50, 200x).

CALRmut

€218,00 plus VAT

#DIA-CAL-100
Anti-CALRmut (Hu) aus Maus (Klon CAL2) – 100 µl
Kategorie:

Beschreibung

Prod.Nr. DIA-CAL-100
Spezifität CALRETICULIN mutation / CALRmut
Speziesreaktivität Human
Speziesreaktivität Mouse
Isotyp IgG2a
Klon CAL2
Anwendung
Immunohistochemistry (IHC),
Formalin-fixed Paraffin-embedded sections (FFPE)
Western Blot
Konjugation unconjugated
Verdünnung IHC 1:50-1:100
Format
Lyophilisate,
Antibody purified (from culture supernatant)
Anzahl 100µl
Control tissue Megakaryocytes from CALRmutated PMNs
Format PBS pH 7.4, with 2% BSA, 0.05% NaN3
Anwendungshinweis Research Use Only
Hersteller ONCOdianova
CALRmut background
Application of anti-CALRmut antibody

Reaktivität

Practical implementation:

CAL2 labels the megakaryocytes in myeloproliferative neoplasms (essential thrombocythaemia (ET) and primary myelofibrosis (PMF))  with CALR mutation and enables to distinguish ET and PMF with CALR mutation from polycythemia vera (PV), from CALR mutation negative ET and PMF and from reactive bone marrow.

Staining pattern:

Cytoplasmic staining of megakaryocytes harboring CALR mutation. The CAL2 IHC assay indicates absence of CALR mutation when all megakaryocytes remain unlabeled.

IHC Protokoll

Staining protocols for anti-human CALRmut antibody clone CAL2
Cat.No.: DIA-CAL
Isotype: Mouse IgG2a
Specificity: Human CALRETICULIN mutations
Physical State: Lyophilisate
Manuelles Protokoll – SKT (Schnellkochtopf)
1. Entparaffinierung:
  • 4x Xylol-Bad – je 5min
  • 1x EtOH 100% – 30 sek
  • 1x EtOH 100% – 5min
  • 1x EtOH 90% – 5min
  • 1x EtOH 70% – 5min
  • in Küvette mit A.dest aufbewahren
2. Hitze-induzierte Epitop-Demaskierung:
Schnellkochtopf /Küvette mit Citratpuffer /10mMol pH6
  • Inkubationszeit – 20min
  • Abkühlung bei Raumtemperatur – 20min
3. Färbung:
  • Auftragen des Primärantikörpers (PAK) CAL2 Verdünnung 1:100
  • 100 μl PAK, Inkubationszeit 30 min
  • in PBS Puffer waschen, 30 sek
  • Auftragen der Sekundärantikörper: Dako REAL™ Detection System, Alkaline Phosphatase/RED
  • Dako REAL™ Link, Biotinylated Sec. AB (AB2) – 30 min
  • in PBS Puffer waschen – 30 sek
  • Dako REAL™ Streptavidin Alk. Phosph. (AP) – 30 min
  • in PBS Puffer waschen – 30 sek
  • Auftragen des Chromogens: Dako REAL™ Chromogen Red
  • 100 μl RED – 16 min
  • in Leitungswasser waschen – 1,5 min
4. Kerngegenfärbung:
  • 100 μl Hämalaun – 7 min
  • in Leitungswasser waschen – 1,5min
4. Eindecken:
  • in Xylol
Automaten Färbeprotokoll – DAKO Omnis:
• Platform:
Dako Omnis
• Kit:
EnVision FLEX HRP Magenta, High pH (Dako Omnis), Code GV900
• Heat-induced epitope retrieval:
EnVision FLEX Target Retrieval Solution, High pH, Tris/EDTA buffer, pH 9 (DM 848), 97 °C, 40 min
• Block:
EnVision FLEX Peroxidase-Blocking Reagent (DM841), 3min
• Secondary antibody (linker):
EnVision FLEX+ Mouse LINKER, 10 min
• Polymer:
EnVision FLEX /HRP (DM842), 20 min
• Chromogen:
EnVision FLEX HRP Magenta Chromogen (DM857), 5min
• Wash buffer:
Wash Buffer, (Dako Omninis, Code GC 807), 10 cycles a 2 min, between each step
• Antibody diluent:
Dako REAL Antibody Diluent (Code S2022)
• Primary antibody:
DIA-CAL, 1:50, 200ul
• Counterstain:

Hematoxylin (Dako Omninis, Code GC808), 7 min

Referenzen

References

  1. Mózes R et al. Calreticulin mutation specific CAL2 immunohistochemistry accurately identifies rare calreticulin mutations in myeloproliferative neoplasms. Pathology, 2018, doi: 10.1016/j.pathol.2018.11.007
  2. Andrici J et al. Mutation specific immuno-histochemistry is highly specific for the presence of calreticulin mutations in myeloproliferative neoplasms. Pathology484: 319-24, 2016.
  3. Nomani L et al. CAL2 Immunohistochemical Staining Accurately Identifies CALR Mutations in Myeloproliferative Neoplasms. Am J Clin Pathol. 1464: 431-438, 2016.
  4. Stein, H et al. A new monoclonal antibody (CAL2) detects CALRETICULIN mutations in formalin-fixed and paraffin embedded bone marrow sections. Leukemia 301: 131-135. 2015.
  5. Nangalia J et al. Somatic CALR Mutations in Myeloproliferative Neoplasms with Nonmutated JAK2. N Engl J Med 369(25): 2391-2405, 2013
  6. Klampfl T et al. Somatic Mutations of Calreticulin in Myeloproliferative Neoplasms. N Engl J Med 369(25): 2379-2390, 2013

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