CD45 Monoclonal Antibody (HI30), Pacific Orange
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Human PBMCs stained with: CD45 Pacific Orange™ (Product # MHCD4530TR), CD4 PerCP-Cy®5.5 (Product # A15858), CD20 APC, CD14 APC-Cy®7 (Product # A15453), CD19 Pacific Green™ (Product # C11210), CD3 Alexa Fluor® 700 (CD0329), HLA-DR PE-Cy®7 (Product # A18558), CD8 Pacific Blue™ (Product # MHCD0828), CD33 FITC (Product # A16185), and CD11c PE (Product # A18674) using the Attune® NxT Acoustic Focusing Cytometer with 405 nm excitation and 440/50 emission filter (Pacific Blue™), 512/25 emission filter (Pacific Green™), and 603/48 emission filter (Pacific Orange™); 488 nm excitation and 530/30 emission filter (FITC) and 695/50 emission filter (PerCP-Cy®5.5); 561 nm excitation and 585/16 emission filter (PE) and 780/60 emission filter (PE-Cy®7); 637 nm excitation and 660/20 emission filter (APC), 720/30 emission filter (Alexa Fluor®700), and 780/60 emission filter (APC-Cy®7). Within the CD3 negative CD45 positive gate, B cells can be identified based on expression of CD19 and CD20. Conventional dendritic cells in peripheral blood are generally negative for T and B cell lineage markers and co-express the integrin CD11c and HLA-DR. Values noted are a percent of the parent CD19 negative CD20 negative gate (top value, no parenthesis) or percent of FSC/SSC lymphocyte gate (bottom value, in parentheses).
Flow Cytometry analysis using a CD45 monoclonal antibody (Product # MHCD4530).
Flow Cytometry analysis using a CD45 monoclonal antibody (Product # MHCD4530).
Figure 2 Specific depletion of human pDC in lymphoid organs in vivo with a human pDC-reactive monoclonal antibody. Humanized Mice were treated with either 15B or isotype control (iso) antibody for 3 times on days -5, -3, -1 prior to termination. Percentages of pDC (Lin - CD4 + CD123 + ) in total human leukocytes (CD45 + ) are analyzed. ( A ) Representative FACS plots and summarized data show relative pDC frequencies before and after antibody treatment in the peripheral blood (n = 7). ( B ) Representative FACS plots and summarized data show pDC depletion by 15B in mesenteric lymph nodes (mLN) and spleen (SP, isotype n = 4; 15B n = 5). All bars in dot graphs indicate median value. Error bars indicate standard deviations (SD). * and ** indicate p<0.05 and p<0.01, respectively. ( C ) Summarized percentages of CD3 - CD19 + B cells in different lymphoid tissues. ( D ) Summarized percentages of CD3 + CD19 - T cells in different lymphoid tissues. ( E ) Summarized percentages of CD3 - CD11c + mDC cells in different lymphoid tissues. All bars in dot graphs indicate median value.
Figure 8 Illustrative example of the immunophenotypic profile of the lymphocyte populations present in normal peripheral blood stained with the lymphoid screening tube (LST) (version 7). ( a ) Typical profile of mature lymphocytes (brown dots) for light scatter parameters and CD45. ( b ) Phenotype of normal mature B-cells for the B-cell-associated markers in the LST combination with a normal distribution according to surface membrane (Sm) light-chain expression (SmIgkappa + B-cells are painted as dark green dots and SmIglamda + B-lymphocytes as light green dots). ( c ) The phenotypic features of normal mature T-cells as defined by the expression of relevant markers in the combination (CD4 + T-cells: dark blue dots; CD8 hi T-cells: blue dots; CD4 - /CD8 -/lo /TCRgammadelta - T-cells: light blue dots; and CD4 - /CD8 -/lo /TCRgammadelta + T-cells: cyan dots). ( d ) Phenotypic pattern of normal peripheral blood NK-cells (yellow dots) for SSC, CD56, CD19/TCRgammadelta, SmCD3, CD38, CD5, CD8 and CD20/CD4 with version 7 of the LST.
Figure 15 An illustration of the strategy used to gate B cell precursor (BCP)-acute lymphoblastic leukemia (ALL) blasts ( a ) and to evaluate their whole immunophenotypic profile using a band dot plot from the Infinicyt software ( b ). The BCP-ALL blast cell population is depicted as blue dots, while normal residual B- and T-cells are shown as purple and green dots, respectively.
Figure 12 Example of a bone marrow (BM) sample from a monoclonal gammopathy of undetermined significance (MGUS) patient stained with the final version (version 6) of the PCD EuroFlow panel illustrating its power for the identification of plasma cells and discrimination between their normal/polyclonal and clonal counterparts. Normal plasma cells (green dots) show a typically normal immunophenotypic profile and coexist in this sample with a clonal population of plasma cells (red dots), which show multiple aberrant phenotypes--CD38 lo , CD45 - , CD19 - , CD56 hi , CD117 + , CD81 lo , CD28 lo and CD27 - --together with high expression of beta2 microglobulin. The polyclonal versus (mono)clonal nature of both plasma cell populations is confirmed by their pattern of expression of cytoplasmic immunoglobulin (CyIg) kappa + and CyIglamda - (normal CyIgkappa/CyIglamda ratio versus CyIglamda + restricted expression, respectively).
Figure 10 EuroFlow small sample tube (SST) analysis of a cerebrospinal fluid (CSF) ( a ) and a vitreous biopsy ( b ) sample with a normal composition of B- and T-lymphocytes. Based on a FSC/SSC/CD45 gating strategy, CD19 + B-cell and SmCD3 + T-cell populations are identified. Even though the surface membrane immunoglobulin (SmIg) kappa and lamda markers are both present in combination with other antibodies with the same fluorochrome, SmIgkappa + (green dots) and SmIglamda + (purple dots) cells can be discerned by gating on the CD19 + B-cell population. In both samples the SmIgkappa + and SmIglamda + B-lymphocytes show a normal ratio (1.5). In a similar way, a normal distribution of CD4 + (orange dots) and CD8 + (blue dots) T-lymphocytes (ratio 2.3 and 2.1 in a and b , respectively) was detected within the CD3 + T-cell population.
Figure 11 EuroFlow small sample tube (SST) analysis of a vitreous biopsy with prominent clonal B-cell population ( a ) and of two cerebrospinal fluid (CSF) samples with prominent aberrant T-cell ( b ) and plasma cell ( c ) populations. In a , following FSC/SSC/CD45 gating, B- and T-cell populations are identified; the B-cell population shows a heavily skewed surface membrane immunoglobulin (SmIg) kappa/SmIglamda ratio (>10), in line with an aberrant, monoclonal large B-lymphocyte population (red dots); residual CD4 + (orange dots) and CD8 + (blue dots) T-lymphocytes show a normal distribution (ratio 2.5). In b , upon FSC/SSC/CD45 gating, a T-cell population is identified, which consists of CD4 + (orange dots) and CD8 + (blue dots) T-lymphocytes, as well as an aberrant T-cell population (red dots) characterized by a SmCD3 lo /CD4 hi /CD8 lo phenotype; the presence of this aberrant T-cell population could be caused by a contaminating blood cell population, given the fact that neutrophil granulocytes can be discerned based on FSC versus SSC and CD45 versus SSC features. Based on a FSC/SSC/CD45 gating strategy, a T-cell population with a normal CD4/CD8 ratio (1.6) can be identified (but no B-cell population) in c . In addition, in this panel a rather large population of CD19 - /CD3 - cells (red dots) is seen that upon further analysis appeared to be CD38 hi and CD56 - , in keeping with a plasma cell origin; further diagnostic work-up of this patient indeed showed an aberrantly si
Figure 21 Characterization of an acute megakaryoblastic leukemia using tube 7 of the EuroFlow acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS) antibody panel. The AML cells (brown and red dots) are positive for CD34 and CD117 and they partly express the megakaryocytic-lineage-associated markers CD42b, CD41 and CD9. HLADR and CD25 were both negative on blast cells.
Figure 20 Identification of basophils (blue events) and plasmocytoid dendritic cells (red events) in a representative bone marrow from a healthy subject using tube 6 of the EuroFlow acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS) antibody panel.
Host / Isotype
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PBS with 4mg/mL BSA
0.1% sodium azide
4° C, store in dark
Product Specific Information
Our Pacific Orange™ dye is optimally excited by the violet laser, and it is recommended that a 575, 585 or 600 nm band pass filter be used for optimal detection. Pacific Blue™ and Pacific Orange™ dye conjugates can be simultaneously excited at 405 nm and emit at 455 nm and 551 nm, respectively, facilitating two-color analysis.
CD45 (LCA, leukocyte common antigen) is a receptor-type protein tyrosine phosphatase (PTP) ubiquitously expressed in all nucleated hematopoietic cells, comprising approximately 10% of all surface proteins in lymphocytes. CD45 is absent on non-hematopoietic cell lines, normal and malignant, non-hematopoietic tissues. CD45 glycoprotein is crucial in lymphocyte development and antigen signaling, serving as an important regulator of Src-family kinases. CD45 protein exists as multiple isoforms as a result of alternative splicing, differ in their extracellular domains but share identical transmembrane and cytoplasmic domains. CD45RA is an isoform of the CD45 complex and has restricted expression between different subtypes of lymphoid cells. CD45 isoforms differ in their ability to translocate into the glycosphingolipid-enriched membrane domains and their expression depends on cell type and physiological state of the cell. CD45 has been shown to be an essential regulator of T- and B-cell antigen receptor signaling and suppresses JAK kinases to regulate cytokine receptor signaling. CD45 is also important in promoting cell survival by modulating integrin-mediated signal transduction pathway, DNA fragmentation during apoptosis and inhibition or upregulation of various immunological functions.
Analyte Specific Reagent
CD45; CD45 antigen; L-CA; Leukocyte common antigen; protein tyrosine phosphatase, receptor type, C; protein tyrosine phosphatase, receptor type, c polypeptide; Receptor-type tyrosine-protein phosphatase C; T200; T200 glycoprotein; T200 leukocyte common antigen; T220 and B220
B220; CD45; CD45R; GP180; L-CA; LCA; LY5; PTPRC; T200
Entrez Gene ID: