Mouse Anti-CD44vRA Recombinant Antibody (clone F8:33) (CAT#: HPAB-0057-YC)

Figure 1 Selective binding of F8:33 anti-CD44vRA mAb to Namalwa cells transfected with CD44vRA (Namalwa-CD44vRA).

(A) Binding of commercial antibodies. Binding of commercial anti-pan and anti-variant CD44 (anti-CD44v6) mAbs to Namalwa-Neo (transfected with empty vector-pcDNA3.1) and Namalwa-CD44s (transfected with standard CD44 cDNA) as well as to Namalwa-CD44v3-v10 and Namalwa-CD44vRA (transfected with CD44v3-v10 and CD44vRA cDNAs, respectively) was detected with fluorescein-labeled anti-mouse Ig (second antibody). First histogram (faint line) in each panel: binding of second antibody only. (B) Binding of anti-CD44vRA mAb: binding of F8:33 anti-CD44vRA to Namalwa transfectants was detected with fluorescein-labeled anti-mouse IgG (second antibody). First histogram in each panel: binding of the second antibody (IIAb) only.

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.

Figure 2 Analysis of F8:33 anti-CD44 mAb by ELISA.

F8:33 anti-CD44vRA mAb (A) and anti-pan-CD44 mAb (Hermes-3) (B) were tested for their ability to bind to microwells coated with soluble CD44vRA, CD44v3-v10, CD44s and CD44-Neo. The binding was detected by a colorimetric assay at optical density of 405 nm. (C) Western blot analysis of CD44s-Fc, CD44v3-v10-Fc and CD44vRA-Fc confirms the molecular mass of these soluble proteins.

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.

Figure 3 Exclusive binding of F8:33 anti-CD44vRA mAb to CD44vRA-positive (detected by PstI digestion) synovial fluid cells from an RA patient.

Binding of F8:33 anti-CD44vRA to synovial fluid cells from an RA patient and primary keratinocytes derived from L and M donors. Binding of F8:33 anti-CD44vRA mAb was detected with fluorescein-labeled anti-mouse-Ig antibody (second antibody). The faint histogram depicts binding of second antibody (II Ab) only. A representative finding of samples from at least five patients.

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.

Figure 4 Anti-CD44vRA mAb discriminates between synovial fluid cells from rheumatoid arthritis and osteoarthritis patients.

Binding of different concentrations of F8:33 anti-CD44vRA mAbs to synovial fluid cells isolated from the joints of RA or OA patients was detected by fluorescein-labeled anti-mouse Ig (second antibody). The faint histogram in each panel depicts binding of second antibody only.

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.

Figure 5 Anti-CD44vRA mAbs bind to synovial fluid cells of an RA patient but not to PBLs from the same patient.

The binding of anti-pan-hCD44 mAb, anti-hCD44v6 mAb and anti-CD44vRA mAbs from two clones (F8:33 and MFI-16-11) to synovial fluid cells (SFCs) and peripheral blood leukocytes (PBLs) of a rheumatoid arthritis (RA) patient as well as to synovial fluid cells of an osteoarthritis (OA) patient was detected with fluorescein-labeled anti-mouse Ig (second antibody). The faint histogram in each panel depicts binding of second antibody only. A representative finding of samples from at least 30 patients.

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.

Figure 6 Anti-CD44vRA mAb-induced apoptosis in synovial fluid cells (SFCs) from a rheumatoid arthritis patient, but not in peripheral blood leukocytes (PBLs) of the same patient.

The apoptosis-inducing capacity of F8:33 anti-CD44vRA mAb and anti-pan-hCD44 mAb was analyzed by two-dimensional flow cytometry, using annexin V (x-axis) and propidium iodide (y-axis). (A) Two-dimensional flow cytometry analysis of synovial fluid cells and PBL from an RA patient. Events accumulating at the bottom left hand side quadrate simulate surviving cells. Events accumulating at the bottom and top right hand side quadrates simulate cells in early and late apoptosis, respectively. (B) Graphical description of two-dimensional flow cytometry analysis of apoptosis in synovial fluid cells and PBL from a different RA patient. A representative finding of samples from at least 30 patients.

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.

Figure 7 Anti-CD44vRA enhanced apoptosis in synovial fluid cells of a rheumatoid arthritis (RA) patient but not, or less, in corresponding cells of two osteoarthritis (OA) patients.

The apoptotic-inducing capacity of F8:33 anti-CD44vRA mAb and anti-pan-hCD44 mAb was analyzed by two-dimensional flow cytometry

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.

Figure 8 Anti-human CD44vRA mAbs reduce joint inflammation in mice with collagen-induced arthritis (CIA).

KM81 anti-mouse pan CD44 mAb and anti-human CD44vRA mAbs from two clones (F8:33, MF-1-16-11) were injected into DBA/1 mice with CIA (see Section 2) at disease onset and then every other day for 12 days. Isotype-matched anti-mouse mAb (4D2; five mice) or anti-human mAb (Serotec nonfunctional anti-CD44 mAb; five mice) were injected, using the same experimental protocol, into a control group. Joint inflammation was assessed by measuring footpad swelling with a micro-caliper. *, P < 0.05; **, P < 0.01, by Student's t-test for unpaired values when compared with the control mouse group. n = Number of mice in each group.

Golan, I., Nedvetzki, S., Golan, I., Eshkar-Sebban, L., Levartovsky, D., Elkayam, O.,... & Naor, D. (2007). Expression of extra trinucleotide in CD44 variant of rheumatoid arthritis patients allows generation of disease-specific monoclonal antibody. Journal of autoimmunity, 28(2-3), 99-113.