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GAD2/GAD65 Overexpression Lysate

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Western Blot: GAD65 Overexpression Lysate (Adult Normal) [NBP2-11101] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for GAD65.

Product Details

Summary
Reactivity HuSpecies Glossary
Applications WB

Order Details

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GAD2/GAD65 Overexpression Lysate Summary

Description

GAD2/GAD65 Transient Overexpression Lysate


Expression Host: HEK293T

Plasmid: RC225984

Accession#: NM_001134366

Protein Tag: C-MYC/DDK

You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.
Gene
GAD2

Applications/Dilutions

Dilutions
  • Western Blot
Application Notes
This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (cat# NBP1-71705) present on the protein construct.

Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.
Theoretical MW
65.2 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Packaging, Storage & Formulations

Storage
Store at -80C. Avoid freeze-thaw cycles.
Buffer
RIPA buffer

Lysate Details for Array

Type
Overexpression

Notes

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

Alternate Names for GAD2/GAD65 Overexpression Lysate

  • EC 4.1.1
  • EC 4.1.1.15
  • GAD2
  • GAD65
  • GAD-65
  • GAD65MGC161605
  • glutamate decarboxylase 2 (pancreatic islets and brain, 65kD)
  • glutamate decarboxylase 2 (pancreatic islets and brain, 65kDa)
  • glutamate decarboxylase 2
  • Glutamate decarboxylase 65 kDa isoform
  • Glutamate decarboxylase-2 (pancreas)
  • MGC161607,65 kDa glutamic acid decarboxylase

Background

Antibodies to glutamic acid decarboxylase-65 (GAD65) are present in autoimmune disorders such as insulin-dependent (type 1) diabetes mellitus (IDDM), stiff man syndrome and polyendocrine autoimmune disease. Auto-antibodies to GAD65 are present in 60-70% of individuals with newly diagnosed IDDM and thus are important markers for disease activity. These auto-antibodies usually recognize conformation-dependent regions on GAD65 and rarely bind to the second isoform, GAD67. Auto-antibodies to GAD67 are found in only 15% of recent-onset IDDM patients and most of this binding can be blocked with GAD65, suggesting shared epitopes between the two isoforms of GAD.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

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Bioinformatics

Gene Symbol GAD2