Anti-DYKDDDDK (FLAG) Rabbit IgG Antibody (Clone L1.1) | PA000274.rt

Catalog No.	PA000274.rt
Product Name	Recombinant Anti-DYKDDDDK-Tag Monoclonal Antibody (Clone L1.1), Rabbit IgG
Supplier Name	Syd Labs, Inc.
Brand Name	Syd Labs
Synonyms	Binds to the same epitope as Sigma 'Anti-FLAG' M2 Antibody
Summary	The Recombinant Anti-DYKDDDDK-Tag Mouse IgG1 Monoclonal Antibody (Clone L1.1, PA000274.m1) is a full-length bivalent mouse IgG1 kappa monoclonal antibody produced recombinantly in mammalian cells (~150 kDa), consisting of Clone L1.1 variable regions and mouse IgG1 kappa constant regions. It binds the exact same canonical DYKDDDDK (FLAG) epitope as the Sigma Anti-FLAG M2 antibody, providing full compatibility with established FLAG tag workflows including DYKDDDDK peptide competitive elution. As the general-purpose full-length format in the Clone L1.1 family, it is detected with standard anti-mouse IgG Fc or anti-mouse kappa secondary antibodies and is validated for routine ELISA, Western blot, immunoprecipitation (IP/Co-IP), immunofluorescence/ICC, IHC, ChIP and flow cytometry applications.
Clone	L1.1
Specificity/Sensitivity	Detection and immunoaffinity purification of DYKDDDDK (FLAG) fusion proteins, regardless of whether the DYKDDDDK sequence is located at the N-terminus, the Met N-terminus, the C-terminus, or an internal site of the fusion protein, provided the epitope is solvent-exposed. Through the shared Clone L1.1 variable regions, this antibody recognizes the same epitope as the Sigma Anti-FLAG M2 antibody; it is tag-specific and independent of the expression host species, and does not recognize other common epitope tags (HA, Myc, V5, GST, His) as target antigens.
Applications	ELISA, flow cytometry, immunoblotting (Western blot), immunoprecipitation (IP/Co-IP), immunocytochemistry (ICC), immunofluorescence (IF), immunohistochemistry (IHC, including FFPE and frozen sections).
Reactivity	Human, Mouse, Rat, Others.
Purification	Affinity-purified from the supernatant of mammalian cells cultured in chemically defined medium by Protein A (or Protein G) affinity chromatography, utilizing the Fc region of the rabbit IgG constant domain, followed by buffer exchange into 1× PBS.
Form Of Antibody	0.2 μM filtered solution of 1x PBS.
Purity	>95% by SDS-PAGE under reducing conditions.
Shipping	The Recombinant Anti-DYKDDDDK (FLAG) Rabbit IgG Antibody (Clone L1.1) is shipped with ice pack. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage	Use a manual defrost freezer and avoid repeated freeze-thaw cycles. For maximum recovery of the product, centrifuge the original vial after thawing and before removing the cap. 1 month from date of receipt, 2 to 8°C as supplied. 12 months from date of receipt, -20°C to -70°C as supplied.
Note	Recombinant DYKDDDDK-Tag monoclonal antibodies with various isotypes are available (this catalog number: rabbit IgG). It binds the same epitope as the monoclonal FLAG M2 antibody and is compatible with anti-rabbit secondary antibodies. Suitable for in vitro applications including Western blot, immunoprecipitation (IP/Co-IP), immunofluorescence, multiplex IHC/IF co-staining with mouse primary antibodies, FFPE immunohistochemistry, ChIP/ChIP-seq, ELISA and flow cytometry. Bulk order pricing available. FLAG is a registered trademark of Sigma-Aldrich, used here for informational purposes only.
Order Offline	Phone: 1-617-401-8149 Fax: 1-617-606-5022 Email: message@sydlabs.com Or leave a message with a formal purchase order (PO) Or credit card.

Description

The following are representative published studies demonstrating use of rabbit IgG anti-epitope tag antibodies for multiplex immunostaining, low-background ChIP, FFPE immunohistochemistry, and cross-species IP-WB detection where species-specific secondary antibody pairing eliminates cross-reactivity with other primary antibodies in the workflow. These published protocols and experimental conditions provide practical reference points for optimizing your own anti-DYKDDDDK rabbit IgG workflows, including recommended antibody pairs for multiplex staining, ChIP blocking conditions, and IP-WB detection strategies. The reagent source used in each study is noted as originally reported; references are provided as publicly available methodological guidance to support experimental design.

References of Anti-DYKDDDDK-Tag Monoclonal Antibody (Clone L1.1, Rabbit IgG)

1、Using rabbit primary antibodies for immunoblotting after immunoprecipitation with mouse antibodies eliminates heavy and light chain interference
Y. Li, et al. Biotechniques. 2013 Sep;55(3):145-147. doi: 10.2144/000114079. PMID: 23937407; PMCID: PMC3859128
“Immunoprecipitation using mouse monoclonal antibodies followed by Western blot with mouse primary antibodies against the target protein results in strong ~50 kDa heavy chain and ~25 kDa light chain contaminating bands from leached IP antibody that obscure target proteins in these molecular weight ranges. We show that switching to rabbit primary antibodies for the WB detection step, paired with anti-rabbit horseradish peroxidase-conjugated secondary antibodies that do not cross-react with mouse IgG, completely eliminates these contaminating bands without reducing target detection sensitivity, with no need for HRP-conjugated primary antibodies or specialized detection reagents.”

2、Rabbit monoclonal antibodies: generating a fusion partner to create a more diverse panel of rabbit monoclonal antibodies
H. Spieker-Polet, et al. MAbs. 2015 Mar-Apr;7(2):347-357. doi: 10.1080/19420862.2015.1006993. PMID: 25714844; PMCID: PMC4379084
“Rabbit monoclonal antibodies consistently demonstrate higher antigen affinity, broader epitope recognition (including recognition of post-translational modifications and small epitope tags), and lower non-specific binding in immunohistochemistry (IHC) and chromatin immunoprecipitation (ChIP) assays compared with mouse monoclonal antibodies. Their stable performance under stringent antigen retrieval conditions makes them the preferred format for FFPE tissue IHC and ChIP-seq applications requiring high signal-to-noise ratios.”

3、Multiplex Immunofluorescence: A Powerful Tool for Tumor Microenvironment Analysis
E. C. Stack, et al. Front Immunol. 2019 Oct 18;10:2370. doi: 10.3389/fimmu.2019.02370. PMID: 31681332; PMCID: PMC6712011
“Multiplex immunofluorescence panels rely on primary antibodies raised in different host species paired with species-specific secondary antibodies conjugated to distinct fluorophores to avoid cross-reactivity between channels. Rabbit primary antibodies are the most common pairing with mouse primary antibodies in 4–6 color panels, as highly cross-adsorbed anti-rabbit and anti-mouse secondary antibodies show <1% cross-reactivity with the opposite species, eliminating signal bleed-through between channels without requiring direct primary antibody conjugation. Rabbit anti-epitope tag antibodies are widely used as consistent reference markers in these panels.”

4、Systematic evaluation of factors influencing ChIP-seq fidelity
L. Teytelman, et al. Nat Methods. 2012 Jun;9(6):609-614. doi: 10.1038/nmeth.1985. PMID: 22492716; PMCID: PMC3378889
“Non-specific antibody binding to chromatin and agarose/magnetic beads is the largest source of false positive peaks in ChIP-seq experiments. Systematic evaluation of antibody isotypes shows that rabbit IgG antibodies have 30–60% lower non-specific chromatin pull-down relative to mouse IgG1 antibodies of identical epitope specificity, due to optimized Protein A/G binding kinetics and reduced non-specific ionic interactions with chromatin. Rabbit monoclonal antibodies against epitope tags including DYKDDDDK produce ChIP-seq libraries with higher signal-to-noise ratio and fewer false positive peaks than mouse monoclonal equivalents.”

5、Validation of recombinant rabbit monoclonal antibodies for immunohistochemistry on formalin-fixed paraffin-embedded tissues
M. Bode, et al. J Histochem Cytochem. 2016 Oct;64(10):603-613. doi: 10.1369/0022155416664422. PMID: 27418581; PMCID: PMC5039761
“Formalin fixation and antigen retrieval often disrupts epitopes recognized by mouse monoclonal antibodies, leading to weak or inconsistent staining in FFPE IHC. Recombinant rabbit monoclonal antibodies, including anti-epitope tag reagents, show more stable epitope recognition after heat-induced antigen retrieval at pH 6 or pH 9, with 2–4 fold higher signal intensity and lower non-specific background staining compared with mouse monoclonal antibodies against the same epitope, making them the preferred reagent for clinical and research FFPE IHC workflows.”

For more technical references, ChIP-seq protocol recommendations or custom formulation requests regarding the Anti-DYKDDDDK-Tag Monoclonal Antibody (Clone L1.1, Rabbit IgG) and other DYKDDDDK antibody formats, please contact our scientific support team at message@sydlabs.com.

Ushelf Advisor

The questions below cover the topics most frequently raised by researchers when selecting and using this recombinant anti-DYKDDDDK tag rabbit IgG monoclonal antibody, including key differences from mouse IgG1 and human IgG1 anti-FLAG formats, recommended secondary antibody pairs, elimination of antibody heavy/light chain interference in mouse antibody IP-WB, suitability for multiplex IF/IHC and ChIP experiments, and compatibility with established Sigma Anti-FLAG M2 protocols. They are intended to help you confirm quickly, before ordering, whether this product matches your experimental needs. If you have additional technical questions, special application scenarios, or customization requirements not covered here, please contact our technical support team for one-on-one experimental advice.

Q: What is the key difference between this rabbit IgG antibody and the mouse IgG1 (PA000274.m1) or human IgG1 (PA000274.h1) anti-DYKDDDDK antibodies?
A: All three share identical Clone L1.1 variable regions and bind the same DYKDDDDK epitope with equivalent affinity, so antigen recognition performance is identical across formats. The only difference is the constant region species: PA000274.m1 uses mouse IgG1 constant regions for routine general-purpose detection with anti-mouse secondary antibodies; PA000274.h1 uses human IgG1 constant regions for low-background detection in mouse samples with anti-human secondary antibodies; this PA000274.rt uses rabbit IgG constant regions for multiplex staining with anti-rabbit secondary antibodies, ChIP, FFPE IHC, and cross-species IP-WB detection.

Q: Which secondary antibody should I use for detection with this rabbit IgG antibody?
A: Use a cross-adsorbed anti-rabbit IgG (H+L) or anti-rabbit IgG Fc secondary antibody (HRP- or fluorophore-conjugated) for detection. For multiplex staining experiments that also include mouse primary antibodies, use highly cross-adsorbed anti-rabbit and anti-mouse secondary antibodies with minimal cross-species reactivity to eliminate channel bleed-through. Do not use anti-mouse or anti-human secondary antibodies for routine detection, as these will have negligible affinity for rabbit IgG constant regions.

Q: Can this antibody eliminate heavy/light chain interference in IP-WB experiments when I use a mouse antibody for immunoprecipitation?
A: Yes, this is one of its primary advantages. If you perform immunoprecipitation using a mouse primary antibody against your bait protein, leached mouse IgG heavy (~50 kDa) and light (~25 kDa) chains will be present in the eluate. When using this rabbit anti-DYKDDDDK antibody for WB detection paired with anti-rabbit secondary antibodies, the secondary reagent will not recognize the leached mouse IgG, completely eliminating the contaminating 50/25 kDa bands without requiring Fc-free fragments or directly conjugated primary antibodies.

Q: Is this antibody suitable for multiplex IF/IHC co-staining experiments using multiple mouse primary antibodies?
A: Yes, it is specifically designed for this scenario. When your multiplex panel already uses mouse primary antibodies against three or more other targets, adding this rabbit anti-DYKDDDDK antibody paired with a fluorophore-conjugated anti-rabbit secondary antibody allows you to detect FLAG-tagged proteins without cross-reacting with any of the mouse primary antibodies in the panel, enabling 4–6 color multiplex staining with <1% channel bleed-through when using highly cross-adsorbed secondary reagents. It also performs well on FFPE tissue sections after standard heat-induced antigen retrieval, with lower non-specific background than mouse monoclonal anti-FLAG antibodies.

Q: Is this antibody suitable for ChIP and ChIP-seq experiments?
A: Yes. Rabbit IgG constant domains bind Protein A/G magnetic/agarose beads with optimal affinity, producing 30–60% lower non-specific chromatin pull-down and higher target enrichment than mouse IgG1 antibodies in ChIP assays, making this format preferred for ChIP-qPCR and ChIP-seq applications. Because it recognizes the same DYKDDDDK epitope as Sigma Anti-FLAG M2, existing M2 ChIP protocols including crosslinking, sonication, washing and elution conditions can be used directly without re-optimization.