Mouse pan b cell isolation kit 2

Manufactured by Miltenyi Biotec

The Mouse Pan B Cell Isolation Kit II is a laboratory equipment product designed to isolate pan B cells from mouse samples. It enables the separation of B cells from other cell types within the sample.

Automatically generated - may contain errors

Lab products found in correlation

Mouse cd4 t cell isolation kit, by Miltenyi Biotec (2 mentions) Celltrace cfse, by Thermo Fisher Scientific (1 mentions) Facs lsr fortessa flow cytometer, by BD (1 mentions) Bovine serum albumin (bsa), by Serva Electrophoresis (1 mentions) Indo 1, by Thermo Fisher Scientific (1 mentions) Indo 1, by BD (1 mentions) 40 m cell strainer, by BD (1 mentions) Lrs fortessa cytometer, by BD (1 mentions) Rbc lysis buffer, by Thermo Fisher Scientific (1 mentions) Facsaria, by BD (1 mentions) Cytofix cytoperm kit, by BD (1 mentions)

Close spelling variants of this product

MojoSort Mouse Pan B Cell Isolation Kit II Pan B Cell Isolation Kit II B Cell Isolation Kit II Cell isolation kits Isolation Kit II Isolation kits

6 protocols using mouse pan b cell isolation kit 2

Adoptive Transfer of B and CD4+ T Cells

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

B cells and CD4⁺ T cells were isolated from nonexposed or KEL^lo RBC–exposed C57BL/6 recipients. To isolate each population, splenocytes from each group were harvested by sterile mechanical disruption of the spleen and lysis of RBCs. B cells or CD4⁺ T cells were then negatively enriched using a mouse Pan B Cell Isolation Kit II or a mouse CD4⁺ T Cell Isolation Kit (both from Miltenyi Biotec). CD4⁺ T cells or B cells were adoptively transferred into C57BL/6 mice, as done previously (10⁷ CD4⁺ T cells or B cells into C57BL/6 recipients or 5 × 10⁷ B cells into B cell–deficient μMT mice) (25 (link)–30 (link)), followed by KEL RBC transfusion. Following transfer, all recipients were challenged with KEL RBCs. For cellular transfer into μMT mice, recipients were conditioned with gamma radiation treatment (300 rad) 24 h prior to B cell transfer, as done previously (31 (link)).

Arthur C.M., Patel S.R., Smith N.H., Bennett A., Kamili N.A., Mener A., Gerner-Smidt C., Sullivan H.C., Hale J.S., Wieland A., Youngblood B., Zimring J.C., Hendrickson J.E, & Stowell S.R. (2017). Antigen Density Dictates Immune Responsiveness following Red Blood Cell Transfusion. Journal of immunology (Baltimore, Md. : 1950), 198(7), 2671-2680.

+ Open protocol

+ Expand

Adoptive Transfer of CFSE-Labeled B Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

B cells were isolated from spleen homogenate of C57BL/6 mice by using a mouse Pan B Cell Isolation Kit II (130-104-443, Miltenyi Biotec) following the manufacturer’s instructions. Half of the isolated B cells were loaded with 2.5 μM D^bGagL peptide (PanaTecs) (31 (link)) and stained with 1 μM Cell Trace CFSE (Invitrogen) (target cells). The other half of the isolated cells were left unpulsed and stained using a QTracker 655 cell staining kit (Invitrogen) (control cells). Cells (1.5 × 10⁷) of the cell mix or target cells (7.5 × 10⁶) were adoptively transferred to the recipient mice 5 minutes before the beginning of the bone marrow surgery required for the intravital imaging.

Mittermüller D., Otto L., Long Z., Kraus A., Beer A., Hasenberg A., Zelinskyy G., Westmeier J., Hasenkrug K.J., Dittmer U, & Gunzer M. (2023). Regulatory T cells suppress the motility of cytotoxic T cells in Friend retrovirus–infected mice. JCI Insight, 8(13), e167482.

+ Open protocol

+ Expand

Adoptive Transfer of B and T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

For the B cell adoptive transfer experiments, spleens and LNs (inguinal, popliteal, iliac, axillar and brachial) from 5 naïve donor mice per strain (wt or MHCII-/-) were harvested and processed as described above. Cells were pooled for each strain and B cells were isolated by positive selection with the mouse Pan B Cell Isolation Kit II (Miltenyi Biotec) according to the manufacturer's instructions. Approximately 2.5×10⁷ B cells/mouse in 300μl of PBS were transferred into naïve recipient μMT-/- mice by i.v. injection.
For the CD4 T cell adoptive transfer experiments, 5 wt (CD90.2), 5 μMT-/- (CD90.2) and 10 CD90.1 mice were immunized with ID93/GLA-SE. One week later their spleens and dLN (inguinal, popliteal, iliac and axillar) were harvested and processed as described above. Total cells from the 5 wt (CD90.2) and 5 CD90.1 constituting the first combined donor population (control) and 5 μMT-/- (CD90.2) and 5 CD90.1 mice constituting the second combined donor population (experimental) were pooled and CD4 T cells were isolated with the mouse CD4 T Cell Isolation Kit (Miltenyi Biotec) according to the manufacturer's instructions. Approximately 3.5×10⁷ CD4 T cells/mouse in 300μl of PBS were transferred into naïve CD45.1 recipient mice by i.v. injection (see Fig. 5a for a schematic representation of the experimental design).

Cauwelaert N.D., Baldwin S.L., Orr M.T., Desbien A.L., Gage E., Hofmeyer K.A, & Coler R.N. (2016). Antigen presentation by B cells guides programming of memory CD4 T cell responses to a TLR4-agonist containing vaccine. European journal of immunology, 46(12), 2719-2729.

+ Open protocol

+ Expand

Quantifying Intracellular Calcium Mobilization

Check if the same lab product or an alternative is used in the 5 most similar protocols

Multivalent cHEL was generated as previously described by biotinylation of HEL (Ubelhart et al, 2015) and crosslinking with unconjugated streptavidin (Invitrogen) at a ratio of 1:2. Intracellular Ca²⁺ mobilization was measured as described (Storch et al, 2007; Ubelhart et al, 2015). For measurement of Ca²⁺ mobilization cells were loaded with the Ca²⁺‐sensitive dye Indo‐1 (Molecular Probes; Invitrogen) and stimulated with HEL (Sigma) at the indicated concentrations (the amount of HEL was constant in both HEL antigen configurations), BSA (Serva) and 10 μg/ml α‐mouse κLC (polyclonal; Southern Biotech). WT Ramos cells expressing endogenous BCR were stimulated with 10 μg/ml α‐human λLC (polyclonal; Southern Biotech) or 10 μg/ml α‐human μHC antibody (polyclonal; Southern Biotech), respectively. For analysis of Ca²⁺ mobilization in splenocytes, total splenic B cells were pre‐enriched by using the B‐cell isolation kit, mouse or the Pan‐B‐cell isolation kit II, mouse (both from Miltenyi Biotec) for Pten‐deficient cells according to the manufacturer's instructions. To exclude residual non‐B cells, purified cells were stained with α‐CD90.2‐PE (53‐2.1; BD) prior to loading with Indo‐1. Ca²⁺ flux measurements were acquired at a FACS LSR Fortessa flow cytometer (BD).

Setz C.S., Khadour A., Renna V., Iype J., Gentner E., He X., Datta M., Young M., Nitschke L., Wienands J., Maity P.C., Reth M, & Jumaa H. (2019). Pten controls B‐cell responsiveness and germinal center reaction by regulating the expression of IgD BCR. The EMBO Journal, 38(11), e100249.

+ Open protocol

+ Expand

Multiparametric Flow Cytometry of Mouse Splenocytes

Check if the same lab product or an alternative is used in the 5 most similar protocols

Single cell suspensions from mouse spleens were obtained via mechanical disruption through 40 µm cell strainer (Falcon) and red blood cell lysis through 1xRBC Lysis Buffer (eBioscience). Cells were incubated on ice for 30 min with the combination of primary fluorochrome-conjugated antibodies or primary unconjugated biotin antibodies followed by conjugated streptavidin (complete list in Supplementary Table S1) in PBS supplemented with 0.5% BSA. Viability markers were also used according to manufacturer’s instructions. For intracellular isotypes staining, cells were first stained with surface antibodies then fixed and permeabilized with Cytofix/Cytoperm kit (BD Bioscience). Cells were washed, resuspended in PBS containing 2% FCS and acquired on BD LRS Fortessa cytometer (Beckton Dickinson). In cell sorting experiments, total spleen B cells were first purified with the Pan B Cell Isolation Kit II mouse (Miltenyi) following manufacturer’s instructions then sorted into GL7⁺PNA⁺EYFP⁺ GC or GL7^-PNA^-EYFP⁺ memory cells with a FACSAria (Beckton Dickinson). Analyses were performed with FlowJo (Tree Star Inc.) software.

Valeri V., Sochon A., Ye C., Mao X., Lecoeuche D., Fillatreau S., Weill J.C., Reynaud C.A, & Hao Y. (2022). B cell intrinsic and extrinsic factors impacting memory recall responses to SRBC challenge. Frontiers in Immunology, 13, 873886.

+ Open protocol

+ Expand

Tracking B Cell Responses in Mice

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total splenocytes from AID-Cre-EYFP naive or SRBC-primed donor mice were purified through the use of the Pan B Cell Isolation Kit II mouse (Miltenyi) according to manufacturer’s instructions. Total splenocytes from wt CD45.2 naive or SRBC-primed donor mice were purified through the use of the CD4⁺ T Cell Isolation Kit mouse (Miltenyi). Where indicated, GC-depleted B cells were obtained by coupling the Germinal Center B Cell (PNA) MicroBead Kit mouse (Miltenyi) and biotin-conjugated GL7 with anti-biotin MicroBeads. IgD-biotin and anti-biotin MicroBeads mediated depletion was used before cell sorting and cell transfer in somatic mutation analysis experiments. 10x10⁶ donor cells were injected intravenously in sterile conditions in 150 µl PBS into wt CD45.1 naive or SRBC-primed mice. Recipient mice were challenged with SRBCs 2h after cell transfer and, where indicated, were fed with tamoxifen one day after. Spleens were harvested and analyzed 5 days after cell transfer.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!