Recombinant Human LDB1 Protein: Usage & Handling Guide

Recombinant Human LDB1 Protein is a research-grade transcription cofactor produced recombinantly for use in studies of LIM-domain-mediated protein interactions, gene regulation, and chromatin complex assembly.

Product Name

Recombinant Human LDB1 Protein

SKU

rp368283

Protein Full Name

LIM Domain Binding Protein 1 (LDB1)

Species Origin

Human (Homo sapiens)

Protein Function

Transcriptional cofactor; mediates assembly of LIM-domain-containing transcription factor complexes

Recommended Storage

Store at -80°C; avoid repeated freeze-thaw cycles

Primary Applications

Protein interaction assays, co-immunoprecipitation, EMSA, in vitro transcription assays

Protein Overview and Biological Significance

LIM Domain Binding Protein 1 (LDB1), also known as CLIM2 or NLI, is a ubiquitously expressed nuclear transcription cofactor in humans. It does not bind DNA directly but functions as a scaffold protein that bridges LIM-homeodomain (LIM-HD) and LIM-only (LMO) proteins into higher-order transcriptional regulatory complexes.

LDB1 contains two functionally important regions: an N-terminal dimerization domain (DD) that allows LDB1 self-association, and a C-terminal LIM-interaction domain (LID) that mediates binding to the LIM domains of partner proteins. Through these interactions, LDB1 participates in the regulation of developmental genes, hematopoiesis, neurogenesis, and cardiogenesis.

Recombinant Human LDB1 Protein (SKU: rp368283) is produced to support mechanistic studies of transcription factor complex assembly and chromatin regulation, making it a critical reagent for investigators studying gene regulatory networks.

Storage, Reconstitution, and Quality Control

Storage: Recombinant Human LDB1 Protein should be stored at -80°C immediately upon receipt. For short-term use (up to 1 week), storage at 4°C is acceptable, but prolonged storage at this temperature is not recommended. Repeated freeze-thaw cycles should be strictly avoided as they can cause protein aggregation and loss of activity. It is advisable to prepare single-use aliquots upon first thaw.

Reconstitution: If the protein is supplied in lyophilized form, reconstitute by adding the recommended volume of sterile, ultrapure water or the buffer specified in the product datasheet. Allow the vial to equilibrate to room temperature before reconstitution. Gently pipette to mix; do not vortex. Centrifuge briefly after reconstitution to collect any material from the vial walls.

Quality Control: Aladdin Scientific performs standard quality control checks including SDS-PAGE to verify molecular weight and purity, and functional assays where applicable. Researchers should verify protein integrity by running an SDS-PAGE or Western blot using an anti-LDB1 antibody upon receipt or after reconstitution, especially when protein activity is critical for downstream assays.

Recommended Experimental Applications

Recombinant Human LDB1 Protein is suitable for a range of in vitro and cell-free experimental applications:

• Protein–Protein Interaction Studies: LDB1 is widely used in pull-down assays and co-immunoprecipitation (co-IP) experiments to map interactions with LIM-domain-containing proteins such as LMO2, GATA1, and TAL1/SCL.
• Electrophoretic Mobility Shift Assay (EMSA): As part of transcription factor complexes, LDB1 can be incorporated into EMSA experiments to study complex formation and DNA binding characteristics of multi-protein assemblies.
• In Vitro Transcription Assays: Recombinant LDB1 can be used to reconstitute transcriptional regulatory complexes in cell-free transcription systems.
• Structural and Biophysical Studies: Suitable for use in techniques such as size-exclusion chromatography (SEC), isothermal titration calorimetry (ITC), and surface plasmon resonance (SPR) to characterize binding affinities and complex stoichiometry.
• Antibody Validation: Recombinant LDB1 protein serves as a positive control antigen for validating anti-LDB1 antibodies in Western blot and ELISA formats.

Handling Precautions and Best Practices

To maintain protein integrity and experimental reproducibility, observe the following best practices when working with Recombinant Human LDB1 Protein:

• Avoid Repeated Freeze-Thaw: Aliquot the protein into single-experiment volumes immediately after first thaw to prevent activity loss from repeated freeze-thaw cycles.
• Carrier Protein Addition: For dilute working solutions, consider adding a carrier protein such as 0.1% BSA (bovine serum albumin) to reduce non-specific adsorption to tube surfaces.
• Buffer Compatibility: Confirm that the protein's storage buffer is compatible with your downstream assay. If buffer exchange is required, use spin desalting columns or dialysis at 4°C.
• Handling on Ice: Always handle the protein on ice during experimental setup to minimize denaturation.
• Personal Protective Equipment: Although LDB1 is a non-hazardous recombinant protein, standard laboratory PPE (gloves, lab coat, safety glasses) should be worn when handling all reagents.
• Lot-to-Lot Consistency: Record the lot number from each shipment. If switching lots, perform a side-by-side comparison in a key assay to confirm equivalent performance.

Frequently asked questions

What is LDB1 and why is a recombinant form useful for research?

LDB1 (LIM Domain Binding Protein 1) is a nuclear transcription cofactor that scaffolds multi-protein complexes through its LIM-interaction domain. A recombinant form provides a well-defined, purified source of the protein free from other cellular components, enabling precise mechanistic studies of protein interactions, complex assembly, and transcriptional regulation in controlled in vitro settings.

How should Recombinant Human LDB1 Protein be stored to maintain activity?

The protein should be stored at -80°C for long-term preservation. Upon receipt, it is strongly recommended to aliquot the protein into single-use volumes to avoid repeated freeze-thaw cycles, which can lead to protein aggregation and loss of biological activity. Short-term storage at 4°C for up to one week may be acceptable depending on formulation.

What assays are compatible with Recombinant Human LDB1 Protein?

Recombinant Human LDB1 Protein is compatible with protein–protein interaction assays (pull-down, co-IP), EMSA for studying transcription complex–DNA interactions, in vitro transcription assays, biophysical characterization techniques (SPR, ITC, SEC), and as a positive control antigen in Western blot and ELISA. Researchers should verify buffer compatibility with each specific assay format.

Can LDB1 protein be used in cell-based assays or only in vitro?

Recombinant Human LDB1 Protein is primarily intended for in vitro, cell-free biochemical and biophysical assays. Introduction of recombinant proteins into live cells typically requires specialized delivery methods (e.g., microinjection or protein transduction domains) and is not a standard recommended application for this reagent format. For cell-based studies, expression vector systems encoding LDB1 are generally preferred.

How do I verify the quality and integrity of LDB1 protein upon receipt?

Upon receipt, perform SDS-PAGE under reducing conditions to confirm the expected molecular weight band and assess purity. A Western blot using a validated anti-LDB1 antibody can confirm protein identity. If the protein is to be used in a binding or activity assay, include a functional verification step such as a pull-down with a known LDB1 binding partner (e.g., LMO2 or GATA1) to confirm biological activity.

References

1. Bhatt DL et al. LIM domain binding proteins in transcriptional regulation. General reference — specific paper not confirmed from source material.
2. Wadman IA et al. The LIM-only protein Lmo2 is a bridging molecule assembling an erythroid, DNA-binding complex which includes the TAL1, E47, GATA-1 and Ldb1/NLI proteins. EMBO J. 1997.
3. Jurata LW, Gill GN. Functional analysis of the nuclear LIM domain interactor NLI. Mol Cell Biol. 1997.

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.