Protocols   Select Another Resource Report Type

Authors: Kelsey Miller
Group: BioLegend

Proper citation: Kelsey Miller 2016. Intracellular Staining With True-Phos™ Perm Buffer in Cell Suspensions. protocols.io https://dx.doi.org/10.17504/protocols.io.e2fbgbn Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: The Anti-Neu5Gc Antibody Kit contains the essential monospecific polyclonal chicken IgY antibody, along with a negative control primary antibody to detect the presence of Neu5Gc on glycoconjugates by Western blot (WB). Samples to be evaluated are first subjected to SDS-PAGE, followed by transfer to a nitrocellulose or polyvinylidenedifluoride (PVDF) membrane. The membrane is then incubated with affinity-purified polyclonal anti-Neu5Gc to determine the presence of Neu5Gc on the protein of interest.The antibody provided in this kit has been shown to identify as little as 5 pmol of Neu5Gc per ug glycoprotein, which is at or below the current detection limit for conventional analysis by acid release, purification, DMB derivatization, HPLC, and electrospray mass-spectrometry. The Western blot provides additional information in that it confirms that Neu5G is directly linked to the glycoprotein of interest rather than to an accompanying sample component.

Proper citation: Kelsey Miller 2016. Anti-Neu5Gc Antibody Kit Protocol - Western blot. protocols.io https://dx.doi.org/10.17504/protocols.io.e28bghw Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: This protocol has been optimized to remove washing steps after antibody cocktail and nanobeads incubations, resulting in a shorter and more convenient protocol. This procedure is optimized for the isolation of 107 to 2 x 108 cells per tube from human peripheral blood mononuclear cells (PBMCs). If working with fewer cells keep volumes as indicated for 107 cells. For best results, optimize the conditions to your specific cell number and tissue. Prepare fresh MojoSort™ Buffer solution by diluting the 5X concentrate with sterile distilled water. To maximize the purity of CD4+ T cells we recommend the depletion of monocytes, as human monocytes express CD4. To deplete monocytes from human PBMCs, we recommend the use of MojoSort™ Human CD14 Selection Kit (Cat. No. 480025/480026). Alternatively, you may consider the Human CD4 T Cell Selection Kit (Cat. No. 480038). After monocyte depletion proceed to isolate the CD4+ T cells. You can also use other methods to deplete monocytes. If you wish to include monocytes in your CD4+ cell isolation, depletion is not required

Proper citation: Kelsey Miller 2016. MojoSort™ Human CD4 Nanobeads No Wash Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e3ibgke Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend

Proper citation: Kelsey Miller 2016. Immunofluorescence Microscopy Protocol with Methanol Fixed Cells. protocols.io https://dx.doi.org/10.17504/protocols.io.fkbbksn Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: The cells targeted by the Nanobeads are either selected or depleted by incubating your sample with the directly conjugated magnetic particles. The magnetically labeled fraction is retained by the use of a magnetic separator. After collection of the targeted cells, downstream applications include functional assays, gene expression, phenotypic characterization, etc.

Proper citation: Kelsey Miller 2016. MojoSort™ Nanobeads No Wash Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e2tbgen Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: This procedure is optimized for the isolation of 107 to 2 x 108 cells per tube. If working with fewer than 107 cells, keep volumes as indicated for 107 cells. For best results, optimize the conditions to your specific cell number and tissue. Prepare fresh MojoSort™ Buffer solution by diluting the 5X concentrate with sterile distilled water. The purity of the CD3- CD49b+ NK Cell fraction typically ranges from 80 to 90%. Removal of dead cells is highly recommended prior to cell isolation. Dead cells can be removed using appropriate centrifugation media or other methods.Product description and procedure summary: The kit is designed for the isolation of mouse NK cells from lymphoid tissues. Target cells are depleted by incubating the sample with the biotin antibody cocktail followed by incubation with magnetic Streptavidin Nanobeads. The magnetically labeled fraction is retained by the use of a magnetic separator. The untouched cells are collected. These are the cells of interest; do not discard the liquid. Some of the downstream applications include functional assays, gene expression, phenotypic characterization, etc.

Proper citation: Kelsey Miller 2016. MojoSort™ Mouse NK Cell Isolation Kit Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.fxpbpmn Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: This protocol is essentially the same as a standard ELISA protocol, with one major exception – all animal compounds must be avoided in buffers, etc. because they contain Neu5Gc, which will interfere with antibody activity and result in misleading data. Essentially, the glycoconjugate to be evaluated is immobilized on an ELISA plate and exposed to a primary antibody. The antibody provided in this kit has been shown to identify as little as 5 pmol of Neu5Gc per microgram glycoprotein, which is at or below the current detection limit for conventional DMB HPLC analysis. An advantage of the ELISA procedure is that it confirms the presence of Neu5Gc on the glycoconjugate of interest, but use caution, as it also reacts with solution/media that contains glycoconjugate.

Proper citation: Kelsey Miller 2016. Anti-Neu5Gc Antibody Kit Protocol - ELISA. protocols.io https://dx.doi.org/10.17504/protocols.io.e27bghn Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend

Proper citation: Kelsey Miller 2016. Alpha-Synuclein ELISA Kit Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e34bgqw Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: Using UV-induced peptide exchange, MHC/peptide monomers can be generated with conditional Flex-T™ monomers that harbor peptides of interest in their binding grooves. These new MHC monomers are subsequently multimerized using streptavidin-fluorophore conjugates. The resulting Flex-T™ reagents can be used for staining antigen-specific T cells and flow cytometric analysis. In humans, the MHC molecules are called HLA (Human Leukocyte Antigen).

Proper citation: Kelsey Miller 2016. Flex-T™ Tetramer and Cell Staining Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e7bbhin Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: The Anti-Neu5Gc Antibody Kit contains the essential monospecific polyclonal chicken IgY antibody, along with a negative control primary antibody to detect the presence of Neu5Gc on glycoconjugates by Western blot (WB). Samples to be evaluated are first subjected to SDS-PAGE, followed by transfer to a nitrocellulose or polyvinylidenedifluoride (PVDF) membrane. The membrane is then incubated with affinity-purified polyclonal anti-Neu5Gc to determine the presence of Neu5Gc on the protein of interest.The antibody provided in this kit has been shown to identify as little as 5 pmol of Neu5Gc per ug glycoprotein, which is at or below the current detection limit for conventional analysis by acid release, purification, DMB derivatization, HPLC, and electrospray mass-spectrometry. The Western blot provides additional information in that it confirms that Neu5G is directly linked to the glycoprotein of interest rather than to an accompanying sample component.

Proper citation: Kelsey Miller 2017. Anti-Neu5Gc Antibody Kit Protocol - Western Blot. protocols.io https://dx.doi.org/10.17504/protocols.io.hvrb656 Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend

Proper citation: Kelsey Miller 2016. Propidium Iodide Cell Cycle Staining Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e2mbgc6 Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: This protocol is essentially the same as a standard ELISA protocol, with one major exception – all animal compounds must be avoided in buffers, etc. because they contain Neu5Gc, which will interfere with antibody activity and result in misleading data. Essentially, the glycoconjugate to be evaluated is immobilized on an ELISA plate and exposed to a primary antibody. The antibody provided in this kit has been shown to identify as little as 5 pmol of Neu5Gc per microgram glycoprotein, which is at or below the current detection limit for conventional DMB HPLC analysis. An advantage of the ELISA procedure is that it confirms the presence of Neu5Gc on the glycoconjugate of interest, but use caution, as it also reacts with solution/media that contains glycoconjugate.

Proper citation: Kelsey Miller 2017. Anti-Neu5Gc Antibody Kit Protocol - ELISA. protocols.io https://dx.doi.org/10.17504/protocols.io.hvpb65n Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: Target cells are depleted by incubating your sample with the biotin antibody cocktail followed by incubation with magnetic Streptavidin Nanobeads (Cat. No. 480015/480016). The magnetically labeled fraction is retained by the use of a magnetic separator. The untouched cells are collected. These are your cells of interest; do not discard the liquid. Some of the downstream applications include functional assays, gene expression, phenotypic characterization, etc.

Proper citation: Kelsey Miller 2016. MojoSort™ Isolation Kits No Wash Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e2wbgfe Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend

Proper citation: Kelsey Miller 2016. Immunohistochemistry Protocol for Sternberger Monoclonal Antibodies. protocols.io https://dx.doi.org/10.17504/protocols.io.e2jbgcn Copy   

•   Source:

Authors: Kelsey Miller, Sam Li
Group: BioLegend
Summary: BioLegend MojoSort™ nanobeads work in commonly used separation columns, based on our internal research as well as validation by external testing by academic labs. This simple protocol consists of following the MojoSort™ protocol to label the cells with pre-diluted MojoSort™ reagents and using the columns as indicated by the manufacturer.Note: Due to the properties of our beads, it may be possible to use far fewer beads and less antibody cocktail that with other commercial suppliers. We recommend a titration to find the best dilution factor. However, as a general rule, dilutions ranging from 1:2 to 1:10 for the antibody cocktail can be used. Dilutions ranging from 1:5 to 1:20 for the Streptavidin Nanobeads can be used. Please contact BioLegend Technical Service ([email protected]) if further assistance is needed.

Proper citation: Kelsey Miller, Sam Li 2019. MojoSort™ Streptavidin Nanobeads Column Protocol - Negative Selection. protocols.io https://dx.doi.org/10.17504/protocols.io.693hh8n Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: The cells targeted by the Nanobeads are either selected or depleted by incubating your sample with the directly conjugated magnetic particles. The magnetically labeled fraction is retained by the use of a magnetic separator. After collection of the targeted cells, downstream applications include functional assays, gene expression, phenotypic characterization, etc.Note: For Human CD4 Nanobeads, please refer to the protocol found on the product webpage for MojoSort™ Human CD4 Nanobeads (Cat. No. 480013/480014).

Proper citation: Kelsey Miller 2016. MojoSort™ Nanobeads Regular Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e2sbgee Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: The cells targeted by the Streptavidin Nanobeads are either selected or depleted by incubating your sample with the magnetic particles after incubating with a biotin-conjugated antibody or antibody cocktail. The magnetically labeled fraction is retained by the use of a magnetic separator. After collection of the targeted cells, downstream applications include functional assays, gene expression, phenotypic characterization, etc.

Proper citation: Kelsey Miller 2016. MojoSort™ Streptavidin Nanobeads Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e2rbgd6 Copy   

•   Source:

Authors: Kelsey Miller, Sam Li
Group: BioLegend
Summary: BioLegend MojoSort™ nanobeads work in commonly used separation columns, based on our internal research as well as validation by external testing by academic labs. This simple protocol consists of following the MojoSort™ protocol to label the cells with pre-diluted MojoSort™ reagents and using the columns as indicated by the manufacturer.Note: Due to the properties of our beads, it may be possible to use far fewer beads and less antibody than with other commercial suppliers. We recommend a titration to find the best dilution factor. However, as a general rule, dilutions ranging from 1:3 to 1:20 for the Nanobeads can be used. Please contact BioLegend Technical Service ([email protected]) if further assistance is needed.

Proper citation: Kelsey Miller, Sam Li 2019. MojoSort™ Streptavidin Nanobeads Column Protocol - Positive Selection. protocols.io https://dx.doi.org/10.17504/protocols.io.692hh8e Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend

Proper citation: Kelsey Miller 2016. Intracellular Staining With True-Phos™ Perm Buffer in Whole Blood. protocols.io https://dx.doi.org/10.17504/protocols.io.e2dbga6 Copy   

•   Source:

Authors: Kelsey Miller
Group: BioLegend
Summary: Cell Fixation and Permeabilization Protocol Using 70% Ethanol

Proper citation: Kelsey Miller 2016. Ethanol Fixation Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.e24bggw Copy   

•   Source: