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  1. Link

    products/primary-antibodies/lysozyme-antibody-epr29942-ab108508.pdf

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Immunology Innate Immunity Macrophage / Inflamm.
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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

  • Datasheet
  • SDS
Reviews (4)Q&A (4)References (81)

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Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunocytochemistry/ Immunofluorescence - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
  • Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR2994(2)] to Lysozyme
  • Suitable for: ICC/IF, WB, IHC-P
  • Knockout validated
  • Reacts with: Mouse, Human

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Knockout
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Human LYZ (Lysozyme) knockout THP-1 cell line (ab269595)

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Overview

  • Product name

    Anti-Lysozyme antibody [EPR2994(2)]
    See all Lysozyme primary antibodies
  • Description

    Rabbit monoclonal [EPR2994(2)] to Lysozyme
  • Host species

    Rabbit
  • Tested applications

    Suitable for: ICC/IF, WB, IHC-Pmore details
    Unsuitable for: IP
  • Species reactivity

    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: THP-1, HepG2, RAW 264.7 and HL-60 whole cell lysate; Human spleen tissue lysate; Natural human Lysozyme protein. IHC-P: Human tonsil, spleen, lung, kidney, brain, breast and heart tissues; Mouse spleen and small intestine tissues. ICC/IF: THP-1 cells.
  • General notes

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 40% Glycerol, 0.05% BSA, PBS
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR2994(2)
  • Isotype

    IgG
  • Research areas

    • Immunology
    • Innate Immunity
    • Macrophage / Inflamm.
    • Immunology
    • Adaptive Immunity
    • T Cells
    • Cytotoxic Cells
    • Cardiovascular
    • Blood
    • Other
    • Tags & Cell Markers
    • Subcellular Markers
    • Organelles
    • Lysosome

Associated products

  • Alternative Versions

    • Anti-Lysozyme antibody [EPR2994(2)] - BSA and Azide free (ab185129)
    • Biotin Anti-Lysozyme antibody [EPR2994(2)] (ab197704)
    • HRP Anti-Lysozyme antibody [EPR2994(2)] (ab197705)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • KO cell lines

    • Human LYZ (Lysozyme) knockout THP-1 cell line (ab269595)
  • KO cell lysates

    • Human LYZ (Lysozyme) knockout THP-1 cell lysate (ab270711)
  • Positive Controls

    • HL-60 whole cell lysate (ab7914)
  • Recombinant Protein

    • Recombinant Human Lysozyme protein (ab158839)
  • Related Products

    • Prestained Protein Ladder – Broad molecular weight (10-245 kDa) (ab116028)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab108508 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
ICC/IF
1/250.

 For unpurified use at 1/500 - 1/1000.

WB
1/10000 - 1/50000. Predicted molecular weight: 17 kDa.
IHC-P (4)
1/1000 - 1/5000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Heat up to 98°C, below boiling, and then let cool for 10-20 min.

See IHC antigen retrieval protocols.

Notes
ICC/IF
1/250.

 For unpurified use at 1/500 - 1/1000.

WB
1/10000 - 1/50000. Predicted molecular weight: 17 kDa.
IHC-P
1/1000 - 1/5000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Heat up to 98°C, below boiling, and then let cool for 10-20 min.

See IHC antigen retrieval protocols.

Application notes
Is unsuitable for IP.

Target

  • Function

    Lysozymes have primarily a bacteriolytic function; those in tissues and body fluids are associated with the monocyte-macrophage system and enhance the activity of immunoagents.
  • Involvement in disease

    Amyloidosis 8
  • Sequence similarities

    Belongs to the glycosyl hydrolase 22 family.
  • Cellular localization

    Secreted.
  • Target information above from: UniProt accession P61626 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 4069 Human
    • Entrez Gene: 17105 Mouse
    • Entrez Gene: 17110 Mouse
    • Omim: 153450 Human
    • SwissProt: P61626 Human
    • SwissProt: P08905 Mouse
    • Unigene: 524579 Human
    • Alternative names

      • 1 4 beta N acetylmuramidase C antibody
      • 1 antibody
      • 4-beta-N-acetylmuramidase C antibody
      • EC 3.2.1.17 antibody
      • LYSC_HUMAN antibody
      • Lysosyme antibody
      • Lysozyme (renal amyloidosis) antibody
      • Lysozyme C antibody
      • Lysozyme C precursor antibody
      • LYZ antibody
      • LZM antibody
      • Renal amyloidosis antibody
      see all

    Images

    • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      All lanes : Anti-Lysozyme antibody [EPR2994(2)] (ab108508) at 1/10000 dilution

      Lane 1 : Wild-type THP-1 (Human monocytic leukemia cell line) whole cell lysate
      Lane 2 : LYZ knockout THP-1 (Human monocytic leukemia cell line) whole cell lysate
      Lane 3 : Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate
      Lane 4 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 17 kDa
      Observed band size: 16 kDa why is the actual band size different from the predicted?



      Lanes 1 - 4: Merged signal (red and green). Green - ab108508 observed at 16 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.

      ab108508 was shown to react with Lysozyme in wild-type THP1 cells in Western blot with loss of signal observed in LYZ knockout sample.Wild-type THP1 and LYZ knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab108508 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 10000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Immunohistochemical analysis of paraffin-embedded human lung tissue sections labeling lysozyme with purified ab108508 at a dilution of 1/1500 (0.6 μg/ml). ab97051 Goat Anti-Rabbit IgG H&L (HRP) at 1/500 was used as the secondary anitbody.Sections were counterstained with Hematoxylin. Antigen retrieval was heat mediated using EDTA Buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

    • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Anti-Lysozyme antibody [EPR2994(2)] (ab108508) at 1/10000 dilution (purified) + RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 20 µg

      Secondary
      Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/2000 dilution

      Predicted band size: 17 kDa
      Observed band size: 16 kDa why is the actual band size different from the predicted?



      Blocking/Diluting buffer 5% NFDM/TBST
    • Immunocytochemistry/ Immunofluorescence - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunocytochemistry/ Immunofluorescence - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Immunocytochemistry/Immunofluorescence analysis of THP-1 (Human monocytic leukemia) cells labeling lysozyme with purified ab108508 at 1/250. Cells were fixed with 100% methanol. ab150077, Alexa Fluor®488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody. Cells were counter-stained with ab195889 Anti-Alpha Tubulin antibody [DM1A] (1/200, 2.5 g/mL) - Microtubule Marker (Alexa Fluor®594) at 1/200. DAPI (blue) was used as a nuclear counterstain. Secondary Only Control: PBS was used instead of the primary antibody as the negative control.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)This image is courtesy of an Abreview submitted by Mr. Thorsten Cramer.

      Paraffin-embedded mouse small intestine tissue stained for Lysozyme using ab108508 in immunohistochemical analysis.

      See Abreview

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Immunohistochemical analysis of paraffin-embedded mouse spleen tissue sections labeling lysozyme with purified ab108508 at a dilution of 1/1500 (0.6 μg/ml). ab97051 Goat Anti-Rabbit IgG H&L (HRP) at 1/500 was used as the secondary anitbody. Sections were counterstained with hematoxylin. Antigen retrieval was heat mediated using EDTA Buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control and is shown in the inset.

    • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      All lanes : Anti-Lysozyme antibody [EPR2994(2)] (ab108508) at 1/10000 dilution (unpurified)

      Lane 1 : HL60 (human promyelocytic leukemia cell line) cell lysate
      Lane 2 : Human spleen lysate

      Lysates/proteins at 10 µg per lane.

      Predicted band size: 17 kDa

    • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Anti-Lysozyme antibody [EPR2994(2)] (ab108508) at 1/50000 dilution (purified) + HL-60 (Human promyelocytic leukemia cell line) whole cell lysate at 20 µg

      Secondary
      Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/2000 dilution

      Predicted band size: 17 kDa
      Observed band size: 16 kDa why is the actual band size different from the predicted?



      Blocking/Diluting buffer 5% NFDM/TBST
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Unpurified ab108508, at 1/1000 dilution, staining Lysozyme in Human tonsil by Immunohistochemistry, Paraffin-embedded tissue.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Western blot - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Anti-Lysozyme antibody [EPR2994(2)] (ab108508) at 1 µg/ml (unpurified) + Native human Lysozyme protein (ab91125) at 0.1 µg

      Secondary
      Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

      Developed using the ECL technique.

      Performed under reducing conditions.

      Predicted band size: 17 kDa


      Exposure time: 4 minutes
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Unpurified ab108508, at 1/1000 dilution, staining Lysozyme in Human spleen by Immunohistochemistry, Paraffin-embedded tissue.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Unpurified ab108508, at 1/1000 dilution, staining Lysozyme in Human kidney by Immunohistochemistry, Paraffin-embedded tissue.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Unpurified ab108508 showing negative staining in Normal human brain tissue.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Unpurified ab108508 showing negative staining in Normal human breast tissue.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

      Unpurified ab108508 showing negative staining in Normal human heart tissue.

      Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

    • Anti-Lysozyme antibody [EPR2994(2)] (ab108508)
      Anti-Lysozyme antibody [EPR2994(2)] (ab108508)

    Protocols

    • Western blot protocols
    • Immunohistochemistry protocols
    • Immunocytochemistry & immunofluorescence protocols

    Click here to view the general protocols

    Datasheets and documents

    • SDS download

    • Datasheet download

      Download

    References (81)

    Publishing research using ab108508? Please let us know so that we can cite the reference in this datasheet.

    ab108508 has been referenced in 81 publications.

    • Centonze M  et al. The Fast Track for Intestinal Tumor Cell Differentiation and In Vitro Intestinal Models by Inorganic Topographic Surfaces. Pharmaceutics 14:N/A (2022). PubMed: 35057113
    • Chiang HY  et al. IL-22 initiates an IL-18-dependent epithelial response circuit to enforce intestinal host defence. Nat Commun 13:874 (2022). PubMed: 35169117
    • Chen B  et al. Identification of macrophages in normal and injured mouse tissues using reporter lines and antibodies. Sci Rep 12:4542 (2022). PubMed: 35296717
    • Mead BE  et al. Screening for modulators of the cellular composition of gut epithelia via organoid models of intestinal stem cell differentiation. Nat Biomed Eng 6:476-494 (2022). PubMed: 35314801
    • Kim S  et al. Tissue extracellular matrix hydrogels as alternatives to Matrigel for culturing gastrointestinal organoids. Nat Commun 13:1692 (2022). PubMed: 35354790
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-8 of 8 Abreviews or Q&A

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Lysozyme antibody [EPR2994(2)]

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Mouse Tissue sections (COLON)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: EDTA
    Permeabilization
    No
    Specification
    COLON
    Blocking step
    BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 37°C
    Fixative
    Paraformaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Feb 25 2022

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Lysozyme antibody [EPR2994(2)]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Mouse Tissue sections (Small intestine)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citrate (pH 6.0)
    Permeabilization
    No
    Specification
    Small intestine
    Blocking step
    Serum as blocking agent for 30 minute(s) · Concentration: 2.5% · Temperature: 25°C
    Fixative
    Formaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    MR. Brett Bell

    Verified customer

    Submitted Dec 28 2020

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Lysozyme antibody [EPR2994(2)]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Rat Tissue sections (Spleen)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citrate pH 6.0
    Permeabilization
    No
    Specification
    Spleen
    Blocking step
    Serum as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 21°C
    Fixative
    Paraformaldehyde
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Abcam user community

    Verified customer

    Submitted Jul 31 2018

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Lysozyme antibody [EPR2994(2)]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Mouse Tissue sections (small intestine)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: citrate
    Permeabilization
    No
    Specification
    small intestine
    Blocking step
    AB block Dako X0590 as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: RT°C
    Fixative
    10% NBF
    Read More
    The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

    Mr. Thorsten Cramer

    Verified customer

    Submitted Aug 18 2016

    Question

    The product data sheet says it reacts with both mouse and human, but what I want to know is if it discriminiates between the mouse M or P lysozyme isoform?

    Read More

    Abcam community

    Verified customer

    Asked on Feb 08 2013

    Answer

    This antibody would not distinguish between the mouse M or P lysozyme isoforms as the immunogen sequence in conserved in both.

    Read More

    Abcam Scientific Support

    Answered on Feb 08 2013

    Question

    I contacted you earlier today for a request
    about antibodies reactive against different lysozyme forms.
    Basically I am working with human lysozyme and its two mutants.the sequence
    of WT lysozyme is given here
    http://www.uniprot.org/uniprot/P61626
    the mutants are
    1) I 74 substituted to T
    2) T 88 substituted to N
    please let me know if any of the monoclonal antibodies to lysozyme: e.g.
    ab91653 or ab108508 are expected to be reactive to all three forms. If you
    think there is such an antibody available please let me know what it was
    raised against.

    Read More

    Abcam community

    Verified customer

    Asked on Aug 29 2012

    Answer

    Thank you for providing that information.

    I have had a look at the two rabbit monoclonal against Lysozyme (ab91653 and ab108508). Both of these antibodies should be able to detect all three forms of the protein (the wild type as well as your two mutants). The immunogen used to raise the ab108508 antibody has been taken from a region N-terminal of your mutations, whilst the immunogen used to raise the antibody ab91653 is from the C-terminal domain. Neither immunogen overlaps any of the mutation sites.

    I hope this information has beenof help. If you have any further questions please do not hesitate to ask.

    Read More

    Abcam Scientific Support

    Answered on Aug 29 2012

    Question

    Please can you send me the IHC protocol for this antibody.

    Read More

    Abcam community

    Verified customer

    Asked on May 30 2012

    Answer


    Thank you for contacting us yesterday.

    I have received further information from the lab regarding the IHC protocol for ab108508. I can
    confirm that 0.01M Sodium Citrate Buffer, pH 6.0 was used for antigen retrieval. Here is a copy
    of the protocol:

    Immunohistochemistry Protocol for Paraffin-embedded Tissues

    1. Solutions and reagents
    1.1. Xylene
    1.2. Ethanol, anhydrous denatured, histological grade (100%, 95%, 70%, 50%)
    1.3. Washing buffer/TBST: 1X TBS/0.1% Tween-20, pH to 7.6.
    1.4. Distilled water (dH2O)
    1.5. Antigen Retrieval Solution: 0.01M Sodium Citrate Buffer, pH 6.0
    To prepare Antigen Retrieval stock solutions:
    10X Stock: Dissolve 29.4 g sodium citrate trisodium salt dehydrate (C 6H 5Na 3O 7 2H2O in 1 liter of dH2O. Add 5mL Tween-20.
    1X Working Solution: Mix 200mL 10X stock with 1800mL dH2O; pH to 6.0
    1.6. 3% Hydrogen Peroxide
    1.7. Blocking Buffer: 10% serum in PBS (serum origin depends on the host of the secondary antibody)
    1.8. Primary Antibody Diluent: 5% serum in PBS (serum origin depends on the host of the
    Secondary antibody)
    1.9. Hematoxylin
    1.10. Permanent Mounting Medium

    2. Protocol
    2.1. Deparaffinization/Rehydration
    2.1.1. Heat slides in an oven at 65 °C for 1 hour.
    2.1.2. De-paraffinize/hydrate using the following series of washes: two Xylene washes (3 min each), followed by two 100% ethanol rinses (3 min each), followed by 95% ethanol, 70% ethanol, 50% ethanol, 30% ethanol, followed by TBST wash for 3 min on a shaker.

    2.2. Antigen Retrieval
    This is recommended Heat Induced Epitope Retrieval (HIER) using Decloaking
    Chamber/Pressure Cooker. Hot water bath or Microwave with temperature sensor can be also used
    (protocol would vary depending on the method used).
    2.2.1. Add 500 ml of dH 2O to Decloaker/Pressure Cooker.
    2.2.2. Immerse slides into staining dish containing Antigen Retrieval Solution. Place staining dish into decloaking chamber.
    2.2.3. Program to run for 30 seconds at 125° C, followed by 10 seconds at 90° C.
    2.2.4. Let it cool down to room temperature (10 – 20 minutes).
    2.2.5. Removes slides and rinse in TBST.
    2.2.6. Proceed to Staining step.

    2.3. Staining
    2.3.1. Wash slides with TBST for 3 min on a shaker.
    2.3.2. Inactivate endogenous peroxidase by covering tissue with 3% hydrogen peroxide for 5 min.
    2.3.3. Wash slides three times with TBST (3 min each on a shaker).
    2.3.4. Block slides with the blocking solution for 1 hour.
    2.3.5. Dilute primary antibody in primary antibody diluent per recommendation on data sheet.
    2.3.6. Apply primary antibody to each section and incubate overnight in the humidified chamber (4 ºC).
    2.3.7. Wash slides three times with TBST (3 min each on a shaker).
    2.3.8. Apply to each section secondary HRP-conjugated anti-rabbit antibody diluted in the blocking solution per manufacturer’s recommendation; incubate for 30 min at room temperature.
    2.3.9. Wash slides three times with TBST (5 min each on a shaker).
    2.3.10. Add freshly prepared DAB substrate to the sections and incubate until stain develops (generally 1 min).
    2.3.11. Rinse sections with water.
    2.3.12. Counterstain with Hematoxylin (generally 10 seconds).
    2.3.13. Rinse sections with water.
    2.3.14. Dehydrate samples using two washes with 100% Ethanol (3 min each), followed by
    two rinses with Xylene (3 min each).
    2.3.15. Mount coverslips on slides using permanent mounting medium.

    I hope this information helps and that the antibody works well. Please do not hesitate to contact us should you have further problems.

    Read More

    Abcam Scientific Support

    Answered on May 30 2012

    Question

    For anti-lysozyme antibody ab91563 and ab108508, Which one will react with human but not cross-react with chicken?

    Read More

    Abcam community

    Verified customer

    Asked on Dec 28 2011

    Answer

    Thank you for contacting us. Based on sequence analysis, immunogen of ab91653 does not share any significant sequence similarity with chicken. For the immunogen of ab108508, it shares 67% identity with chicken Lysozyme C, so it should also be human specific. I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

    Read More

    Abcam Scientific Support

    Answered on Dec 28 2011

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