sequencing grade modified trypsin  (Promega)

 
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    Name:
    Sequencing Grade Modified Trypsin
    Description:
    Cleavage sites C terminal of Arg and Lys Optimal pH 7 9
    Catalog Number:
    v5111
    Price:
    None
    Category:
    Protein Analysis Mass Spectrometry Proteases and Surfactants
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    Structured Review

    Promega sequencing grade modified trypsin
    Strategy of SUMO1 <t>modified</t> sites immunoaffinity purification. (A) Representation of the C-terminal <t>sequence</t> comparison of SUMO1 WT , SUMO1 ΔGG and SUMO1 T95R after <t>trypsin</t> digestion. (B) Immunoblot analysis confirmed the expressions of SUMO1-conjugated proteins in SUMO1 WT and SUMO1 T95R . HEK293T cells were stably transfected with indicated plasmids and SUMO1 ΔGG was chosen as the negative control. Ponceau-S staining was shown as loading control. (C) Schematic overview of His 10 -SUMO1 T95R -modified peptides identification. Stably expressing cell samples transfected with indicated plasmids were subjected to the first concentration of SUMOylated proteins and trypsin digestion, followed by peptide IP to enrich SUMO1-conjugated peptides. diGly featured peptides were subsequently exposed for LC-MS/MS identification. (D) Validation of expression patterns and level of conjugation by immunoblot analysis. His 10 -SUMO1 WT , His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells were lysed and subjected to Ni-NTA pulldown assay. The presence of ubiquitin and ubiquitin-like modifier patterns were confirmed with indicated antibodies. (E) Detection of SUMOylation patterns after transfection of SENP family members. SUMOylated proteins from His 10 -SUMO1 T95R stably expressed cells with EV and Flag-tagged SENP family members transfected were pulled down and analyzed by immunoblot using anti-SUMO1, anti-Flag or anti-Actin antibodies. Ponceau-S staining was shown as loading control. (F) Schematic workflow of the deduction strategy for constructing SUMO1-modified proteome. Venn plot indicated the number of overlapped diGly sites detected in His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells.
    Cleavage sites C terminal of Arg and Lys Optimal pH 7 9
    https://www.bioz.com/result/sequencing grade modified trypsin/product/Promega
    Average 99 stars, based on 198 article reviews
    Price from $9.99 to $1999.99
    sequencing grade modified trypsin - by Bioz Stars, 2020-07
    99/100 stars

    Images

    1) Product Images from "Global Screening of Sentrin-Specific Protease Family Substrates in SUMOylation"

    Article Title: Global Screening of Sentrin-Specific Protease Family Substrates in SUMOylation

    Journal: bioRxiv

    doi: 10.1101/2020.02.25.964072

    Strategy of SUMO1 modified sites immunoaffinity purification. (A) Representation of the C-terminal sequence comparison of SUMO1 WT , SUMO1 ΔGG and SUMO1 T95R after trypsin digestion. (B) Immunoblot analysis confirmed the expressions of SUMO1-conjugated proteins in SUMO1 WT and SUMO1 T95R . HEK293T cells were stably transfected with indicated plasmids and SUMO1 ΔGG was chosen as the negative control. Ponceau-S staining was shown as loading control. (C) Schematic overview of His 10 -SUMO1 T95R -modified peptides identification. Stably expressing cell samples transfected with indicated plasmids were subjected to the first concentration of SUMOylated proteins and trypsin digestion, followed by peptide IP to enrich SUMO1-conjugated peptides. diGly featured peptides were subsequently exposed for LC-MS/MS identification. (D) Validation of expression patterns and level of conjugation by immunoblot analysis. His 10 -SUMO1 WT , His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells were lysed and subjected to Ni-NTA pulldown assay. The presence of ubiquitin and ubiquitin-like modifier patterns were confirmed with indicated antibodies. (E) Detection of SUMOylation patterns after transfection of SENP family members. SUMOylated proteins from His 10 -SUMO1 T95R stably expressed cells with EV and Flag-tagged SENP family members transfected were pulled down and analyzed by immunoblot using anti-SUMO1, anti-Flag or anti-Actin antibodies. Ponceau-S staining was shown as loading control. (F) Schematic workflow of the deduction strategy for constructing SUMO1-modified proteome. Venn plot indicated the number of overlapped diGly sites detected in His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells.
    Figure Legend Snippet: Strategy of SUMO1 modified sites immunoaffinity purification. (A) Representation of the C-terminal sequence comparison of SUMO1 WT , SUMO1 ΔGG and SUMO1 T95R after trypsin digestion. (B) Immunoblot analysis confirmed the expressions of SUMO1-conjugated proteins in SUMO1 WT and SUMO1 T95R . HEK293T cells were stably transfected with indicated plasmids and SUMO1 ΔGG was chosen as the negative control. Ponceau-S staining was shown as loading control. (C) Schematic overview of His 10 -SUMO1 T95R -modified peptides identification. Stably expressing cell samples transfected with indicated plasmids were subjected to the first concentration of SUMOylated proteins and trypsin digestion, followed by peptide IP to enrich SUMO1-conjugated peptides. diGly featured peptides were subsequently exposed for LC-MS/MS identification. (D) Validation of expression patterns and level of conjugation by immunoblot analysis. His 10 -SUMO1 WT , His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells were lysed and subjected to Ni-NTA pulldown assay. The presence of ubiquitin and ubiquitin-like modifier patterns were confirmed with indicated antibodies. (E) Detection of SUMOylation patterns after transfection of SENP family members. SUMOylated proteins from His 10 -SUMO1 T95R stably expressed cells with EV and Flag-tagged SENP family members transfected were pulled down and analyzed by immunoblot using anti-SUMO1, anti-Flag or anti-Actin antibodies. Ponceau-S staining was shown as loading control. (F) Schematic workflow of the deduction strategy for constructing SUMO1-modified proteome. Venn plot indicated the number of overlapped diGly sites detected in His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells.

    Techniques Used: Modification, Immunoaffinity Purification, Sequencing, Stable Transfection, Transfection, Negative Control, Staining, Expressing, Concentration Assay, Liquid Chromatography with Mass Spectroscopy, Conjugation Assay

    Related Articles

    Sequencing:

    Article Title: PhosFox: a bioinformatics tool for peptide-level processing of LC-MS/MS-based phosphoproteomic data
    Article Snippet: .. The proteins were reduced, alkylated and enzymatically digested in-solution with lysyl endopeptidase (7.5 μg/sample, rLys-C Mass Spec Grade, Promega) for 2 hrs, which after the samples were diluted with 7 ml of destilled water, followed by digestion with trypsin (20 μg/sample, Sequencing Grade Modified Trypsin, Promega) for 16 hrs. .. Undigested proteins and cell debris were removed, and the samples were desalted on Sep-Pak Vac RP C18 cartridges (Waters).

    Article Title: Adseverin: A novel cisplatin‐resistant marker in the human bladder cancer cell line HT1376 identified by quantitative proteomic analysis), Adseverin: A novel cisplatin‐resistant marker in the human bladder cancer cell line HT1376 identified by quantitative proteomic analysis
    Article Snippet: .. The protein solutions were diluted 4‐fold with 50 mM ammonium bicarbonate solution and digested with 0.5 μg of sequence‐grade trypsin (Promega, Madison, WI) at 37 °C for 12 h. Digestion was terminated by adding 5% trifluoroacetic acid (TFA), and the tryptic peptides were desalted and concentrated on a self‐prepared STAGE tip ( ). .. Tandem MS (MS/MS) and MRM analysis were performed using the QTRAP 5500 hybrid triple quadrupole/linear ion trap system (AB‐SCIEX; Foster, CA) coupled with Prominence NanoLC (Shimadzu, Kyoto, Japan).

    Article Title: Mutations in the v-Rel Transactivation Domain Indicate Altered Phosphorylation and Identify a Subset of NF-?B-Regulated Cell Death Inhibitors Important for v-Rel Transforming Activity
    Article Snippet: .. Radiolabeled v-Rel protein bands were excised from the membrane and treated with trypsin (10 μg; sequencing grade modified trypsin; Promega Corp., Madison, Wis.) in 50 mM ammonium bicarbonate for 2 h at 37°C, as described ( ). ..

    Article Title: A Receptor for Activated C Kinase Is Part of Messenger Ribonucleoprotein Complexes Associated with PolyA-mRNAs in Neurons
    Article Snippet: .. Gel pieces were then hydrated in a trypsin (sequencing grade, unlyophilized modified; Promega, Madison, WI) solution (12.5 ng/μl) for 45 min on ice. ..

    Article Title: TRIM9 and TRIM67 are new targets in paraneoplastic cerebellar degeneration
    Article Snippet: .. Briefly, proteins were in-gel digested using modified trypsin (sequencing grade, Promega, Charbonnières, France), and resulting peptides were analyzed by online nanoLC-MS/MS (UltiMate 3000 and LTQ-Orbitrap Velos Pro, Thermo Scientific, Grenoble, France). .. Peptides and proteins from different samples were identified, filtered, and compared using Mascot and Proline software (Profi Proteomics, Toulouse, France).

    Article Title: The Apaf-1-binding protein Aven is cleaved by Cathepsin D to unleash its anti-apoptotic potential
    Article Snippet: .. Next, samples were boiled for 5 min, placed on ice for 5 min and digested overnight at 37 °C with 2 μ g trypsin (Promega Sequencing-Grade Modified Trypsin, Promega, Mannheim, Germany). .. Without pre-enrichment for N-terminal peptides, 100 μ l of each sample was acidified with 2 μ l formic acid and applied for LC-MS/MS analysis using a Thermo LTQ-Orbitrap XL mass spectrometer (2.5 μ l of sample was injected for each reading), which was operated as previously described.

    Article Title: Analysis of Serum Total and Free PSA Using Immunoaffinity Depletion Coupled to SRM: Correlation with Clinical Immunoassay Tests
    Article Snippet: .. The resulting protein mixtures were diluted 10-fold with 50 mM NH4 CO3 , pH 8.0, before sequencing grade modified porcine trypsin (Promega) was added at a trypsin:protein ratio of 1:50 (w/w). .. The samples were incubated at 37 °C for 3 h. The tryptically digested samples were then loaded onto 1-mL SPE C18 columns (Supelco, Bellefonte, PA) and washed with 4 mL of 0.1% TFA, 5% acetonitrile.

    Article Title: Coordinated Post-translational Responses of Aquaporins to Abiotic and Nutritional Stimuli in Arabidopsis Roots *
    Article Snippet: .. One of these subsequently used Lys-C (Roche Applied Science) for 4 h at 37 °C and trypsin (Sequencing Grade Modified, Promega, Madison, WI) overnight at 37 °C. .. The other used chymotrypsin (Sequencing Grade Modified, Promega) for 5 h at 25 °C and then AspN (Sequencing Grade Modified, Promega) overnight at 25 °C.

    Modification:

    Article Title: PhosFox: a bioinformatics tool for peptide-level processing of LC-MS/MS-based phosphoproteomic data
    Article Snippet: .. The proteins were reduced, alkylated and enzymatically digested in-solution with lysyl endopeptidase (7.5 μg/sample, rLys-C Mass Spec Grade, Promega) for 2 hrs, which after the samples were diluted with 7 ml of destilled water, followed by digestion with trypsin (20 μg/sample, Sequencing Grade Modified Trypsin, Promega) for 16 hrs. .. Undigested proteins and cell debris were removed, and the samples were desalted on Sep-Pak Vac RP C18 cartridges (Waters).

    Article Title: Mutations in the v-Rel Transactivation Domain Indicate Altered Phosphorylation and Identify a Subset of NF-?B-Regulated Cell Death Inhibitors Important for v-Rel Transforming Activity
    Article Snippet: .. Radiolabeled v-Rel protein bands were excised from the membrane and treated with trypsin (10 μg; sequencing grade modified trypsin; Promega Corp., Madison, Wis.) in 50 mM ammonium bicarbonate for 2 h at 37°C, as described ( ). ..

    Article Title: A Receptor for Activated C Kinase Is Part of Messenger Ribonucleoprotein Complexes Associated with PolyA-mRNAs in Neurons
    Article Snippet: .. Gel pieces were then hydrated in a trypsin (sequencing grade, unlyophilized modified; Promega, Madison, WI) solution (12.5 ng/μl) for 45 min on ice. ..

    Article Title: TRIM9 and TRIM67 are new targets in paraneoplastic cerebellar degeneration
    Article Snippet: .. Briefly, proteins were in-gel digested using modified trypsin (sequencing grade, Promega, Charbonnières, France), and resulting peptides were analyzed by online nanoLC-MS/MS (UltiMate 3000 and LTQ-Orbitrap Velos Pro, Thermo Scientific, Grenoble, France). .. Peptides and proteins from different samples were identified, filtered, and compared using Mascot and Proline software (Profi Proteomics, Toulouse, France).

    Article Title: The Apaf-1-binding protein Aven is cleaved by Cathepsin D to unleash its anti-apoptotic potential
    Article Snippet: .. Next, samples were boiled for 5 min, placed on ice for 5 min and digested overnight at 37 °C with 2 μ g trypsin (Promega Sequencing-Grade Modified Trypsin, Promega, Mannheim, Germany). .. Without pre-enrichment for N-terminal peptides, 100 μ l of each sample was acidified with 2 μ l formic acid and applied for LC-MS/MS analysis using a Thermo LTQ-Orbitrap XL mass spectrometer (2.5 μ l of sample was injected for each reading), which was operated as previously described.

    Article Title: Analysis of Serum Total and Free PSA Using Immunoaffinity Depletion Coupled to SRM: Correlation with Clinical Immunoassay Tests
    Article Snippet: .. The resulting protein mixtures were diluted 10-fold with 50 mM NH4 CO3 , pH 8.0, before sequencing grade modified porcine trypsin (Promega) was added at a trypsin:protein ratio of 1:50 (w/w). .. The samples were incubated at 37 °C for 3 h. The tryptically digested samples were then loaded onto 1-mL SPE C18 columns (Supelco, Bellefonte, PA) and washed with 4 mL of 0.1% TFA, 5% acetonitrile.

    Article Title: Coordinated Post-translational Responses of Aquaporins to Abiotic and Nutritional Stimuli in Arabidopsis Roots *
    Article Snippet: .. One of these subsequently used Lys-C (Roche Applied Science) for 4 h at 37 °C and trypsin (Sequencing Grade Modified, Promega, Madison, WI) overnight at 37 °C. .. The other used chymotrypsin (Sequencing Grade Modified, Promega) for 5 h at 25 °C and then AspN (Sequencing Grade Modified, Promega) overnight at 25 °C.

    Mass Spectrometry:

    Article Title: PhosFox: a bioinformatics tool for peptide-level processing of LC-MS/MS-based phosphoproteomic data
    Article Snippet: .. The proteins were reduced, alkylated and enzymatically digested in-solution with lysyl endopeptidase (7.5 μg/sample, rLys-C Mass Spec Grade, Promega) for 2 hrs, which after the samples were diluted with 7 ml of destilled water, followed by digestion with trypsin (20 μg/sample, Sequencing Grade Modified Trypsin, Promega) for 16 hrs. .. Undigested proteins and cell debris were removed, and the samples were desalted on Sep-Pak Vac RP C18 cartridges (Waters).

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  • About
  • News
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  • Team
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  • 94
    Promega sequencing grade modified trypsin
    Strategy of SUMO1 <t>modified</t> sites immunoaffinity purification. (A) Representation of the C-terminal <t>sequence</t> comparison of SUMO1 WT , SUMO1 ΔGG and SUMO1 T95R after <t>trypsin</t> digestion. (B) Immunoblot analysis confirmed the expressions of SUMO1-conjugated proteins in SUMO1 WT and SUMO1 T95R . HEK293T cells were stably transfected with indicated plasmids and SUMO1 ΔGG was chosen as the negative control. Ponceau-S staining was shown as loading control. (C) Schematic overview of His 10 -SUMO1 T95R -modified peptides identification. Stably expressing cell samples transfected with indicated plasmids were subjected to the first concentration of SUMOylated proteins and trypsin digestion, followed by peptide IP to enrich SUMO1-conjugated peptides. diGly featured peptides were subsequently exposed for LC-MS/MS identification. (D) Validation of expression patterns and level of conjugation by immunoblot analysis. His 10 -SUMO1 WT , His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells were lysed and subjected to Ni-NTA pulldown assay. The presence of ubiquitin and ubiquitin-like modifier patterns were confirmed with indicated antibodies. (E) Detection of SUMOylation patterns after transfection of SENP family members. SUMOylated proteins from His 10 -SUMO1 T95R stably expressed cells with EV and Flag-tagged SENP family members transfected were pulled down and analyzed by immunoblot using anti-SUMO1, anti-Flag or anti-Actin antibodies. Ponceau-S staining was shown as loading control. (F) Schematic workflow of the deduction strategy for constructing SUMO1-modified proteome. Venn plot indicated the number of overlapped diGly sites detected in His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells.
    Sequencing Grade Modified Trypsin, supplied by Promega, used in various techniques. Bioz Stars score: 94/100, based on 198 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sequencing grade modified trypsin/product/Promega
    Average 94 stars, based on 198 article reviews
    Price from $9.99 to $1999.99
    sequencing grade modified trypsin - by Bioz Stars, 2020-07
    94/100 stars
      Buy from Supplier

    92
    Promega two step digestion approach
    Strategy of SUMO1 <t>modified</t> sites immunoaffinity purification. (A) Representation of the C-terminal <t>sequence</t> comparison of SUMO1 WT , SUMO1 ΔGG and SUMO1 T95R after <t>trypsin</t> digestion. (B) Immunoblot analysis confirmed the expressions of SUMO1-conjugated proteins in SUMO1 WT and SUMO1 T95R . HEK293T cells were stably transfected with indicated plasmids and SUMO1 ΔGG was chosen as the negative control. Ponceau-S staining was shown as loading control. (C) Schematic overview of His 10 -SUMO1 T95R -modified peptides identification. Stably expressing cell samples transfected with indicated plasmids were subjected to the first concentration of SUMOylated proteins and trypsin digestion, followed by peptide IP to enrich SUMO1-conjugated peptides. diGly featured peptides were subsequently exposed for LC-MS/MS identification. (D) Validation of expression patterns and level of conjugation by immunoblot analysis. His 10 -SUMO1 WT , His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells were lysed and subjected to Ni-NTA pulldown assay. The presence of ubiquitin and ubiquitin-like modifier patterns were confirmed with indicated antibodies. (E) Detection of SUMOylation patterns after transfection of SENP family members. SUMOylated proteins from His 10 -SUMO1 T95R stably expressed cells with EV and Flag-tagged SENP family members transfected were pulled down and analyzed by immunoblot using anti-SUMO1, anti-Flag or anti-Actin antibodies. Ponceau-S staining was shown as loading control. (F) Schematic workflow of the deduction strategy for constructing SUMO1-modified proteome. Venn plot indicated the number of overlapped diGly sites detected in His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells.
    Two Step Digestion Approach, supplied by Promega, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/two step digestion approach/product/Promega
    Average 92 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    two step digestion approach - by Bioz Stars, 2020-07
    92/100 stars
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    Image Search Results


    Strategy of SUMO1 modified sites immunoaffinity purification. (A) Representation of the C-terminal sequence comparison of SUMO1 WT , SUMO1 ΔGG and SUMO1 T95R after trypsin digestion. (B) Immunoblot analysis confirmed the expressions of SUMO1-conjugated proteins in SUMO1 WT and SUMO1 T95R . HEK293T cells were stably transfected with indicated plasmids and SUMO1 ΔGG was chosen as the negative control. Ponceau-S staining was shown as loading control. (C) Schematic overview of His 10 -SUMO1 T95R -modified peptides identification. Stably expressing cell samples transfected with indicated plasmids were subjected to the first concentration of SUMOylated proteins and trypsin digestion, followed by peptide IP to enrich SUMO1-conjugated peptides. diGly featured peptides were subsequently exposed for LC-MS/MS identification. (D) Validation of expression patterns and level of conjugation by immunoblot analysis. His 10 -SUMO1 WT , His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells were lysed and subjected to Ni-NTA pulldown assay. The presence of ubiquitin and ubiquitin-like modifier patterns were confirmed with indicated antibodies. (E) Detection of SUMOylation patterns after transfection of SENP family members. SUMOylated proteins from His 10 -SUMO1 T95R stably expressed cells with EV and Flag-tagged SENP family members transfected were pulled down and analyzed by immunoblot using anti-SUMO1, anti-Flag or anti-Actin antibodies. Ponceau-S staining was shown as loading control. (F) Schematic workflow of the deduction strategy for constructing SUMO1-modified proteome. Venn plot indicated the number of overlapped diGly sites detected in His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells.

    Journal: bioRxiv

    Article Title: Global Screening of Sentrin-Specific Protease Family Substrates in SUMOylation

    doi: 10.1101/2020.02.25.964072

    Figure Lengend Snippet: Strategy of SUMO1 modified sites immunoaffinity purification. (A) Representation of the C-terminal sequence comparison of SUMO1 WT , SUMO1 ΔGG and SUMO1 T95R after trypsin digestion. (B) Immunoblot analysis confirmed the expressions of SUMO1-conjugated proteins in SUMO1 WT and SUMO1 T95R . HEK293T cells were stably transfected with indicated plasmids and SUMO1 ΔGG was chosen as the negative control. Ponceau-S staining was shown as loading control. (C) Schematic overview of His 10 -SUMO1 T95R -modified peptides identification. Stably expressing cell samples transfected with indicated plasmids were subjected to the first concentration of SUMOylated proteins and trypsin digestion, followed by peptide IP to enrich SUMO1-conjugated peptides. diGly featured peptides were subsequently exposed for LC-MS/MS identification. (D) Validation of expression patterns and level of conjugation by immunoblot analysis. His 10 -SUMO1 WT , His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells were lysed and subjected to Ni-NTA pulldown assay. The presence of ubiquitin and ubiquitin-like modifier patterns were confirmed with indicated antibodies. (E) Detection of SUMOylation patterns after transfection of SENP family members. SUMOylated proteins from His 10 -SUMO1 T95R stably expressed cells with EV and Flag-tagged SENP family members transfected were pulled down and analyzed by immunoblot using anti-SUMO1, anti-Flag or anti-Actin antibodies. Ponceau-S staining was shown as loading control. (F) Schematic workflow of the deduction strategy for constructing SUMO1-modified proteome. Venn plot indicated the number of overlapped diGly sites detected in His 10 -SUMO1 ΔGG , SUMO1 T95R and SUMO1 T95K stable cells.

    Article Snippet: Samples were then washed twice with urea wash buffer (250μl of 8M urea, 100mM Tris pH 7.5), three times with 250 μl of 50mM ammonium bicarbonate (ABC) buffer and digested for 16 hours with sequencing-grade modified trypsin (Promega) in 200 μl ABC buffer at 37 °C (enzyme to protein ratio 1:50).

    Techniques: Modification, Immunoaffinity Purification, Sequencing, Stable Transfection, Transfection, Negative Control, Staining, Expressing, Concentration Assay, Liquid Chromatography with Mass Spectroscopy, Conjugation Assay