News & Featured Publications

Anti-HEK Antibodies with 97% HCP Coverage

June 18, 2024

Looking for HEK antibodies with high coverage?

Applied Biomics’ anti HEK HCP antibody offers 97% coverage of many HEK and human cell lines, including HEK293, CAP, Hela, A549, PER.C6, and MRC-5. Our HEK antibodies detect over 3,000 high molecular weight (HMW) and low molecular weight (LMW) host cell proteins (HCPs), and come in Serum or IgG form.

Our HEK antibodies have been repeatedly tested to ensure over 93% coverage in each quadrant of the 2D Western Blot, including those LMW HCPs. They are ideal for use in 1D/2D Western Blots, ELISA, Immunoprecipitation (IP), or as capture/detection anti-human HCP sources.

Want to test our antibodies first? As a leading provider of HCP antibody coverage services, we can test our antibody against your samples and compare with your in-house or other commercial HCP antibodies.Get 30% off our antibodies when testing with our HCP Antibody Coverage service. Simply contact us and use the code ABO30HCP.

Left: 1D Western Blot – HEK antibody coverage of various HEK and human cell lines
Right: Quadrant coverage of Anti-HEK HCP Antibody against HEK HCP

Why us?

  • High HCP coverage: Each HEK antibody product has passed rigorous 2D Western Blot analysis to ensure overall HCP coverage of 97% and each quadrant coverage > 93%, including HMW and LMW HCPs.
  • Competitive pricing: Same quantity at a lower price than similar antibodies from other vendors.
  • Quality guaranteed: We optimize our products to ensure consistency in performance matching the highest standards.
  • Excellent support: Backed by over 20 years of experience in antibody development and validation, our scientists will address your questions / concerns within 48 hours.

Identify Neurodegenerative Disease Biomarkers with 2D DIGE

May 15, 2024

A recent study published in ACS Chemical Neuroscience used our 2D DIGE and Mass Spectrometry platform to identify potential biomarkers for neurodegenerative disease.

Defects in protein RanBP2 are linked to various aging-related diseases, but the mechanism is not well understood. In the study, a 2D DIGE experiment was performed to compare the proteomic profiles of WT, RanBP2 negative, and RanBP2 defective mouse tissues.

2D DIGE detected 67 spots with significant differential expression among the samples. In particular, three proteins were consistently increased in the RanBP2 defective sample and decreased in the negative sample. These were then identified by Mass Spectrometry as y-crystallins.

Figure 1: Zoomed-in 2D DIGE images of WT vs RanBP2 negative and WT vs RanBP2 defective mouse tissue. Spots 3, 4, and 5 were consistently downregulated in the first comparison and upregulated in the second comparison.

Further validation assays provided insights into the function of these crystallins with respect to RanBP2, and the relative levels of their isoforms. Ultimately, the scientists developed a model in which RanBP2 reshapes crystallins to suppress ubiquitination and slow the aging process.

Comments: The data here showed once again our 2D DIGE/MS platform could reliably identify key biomarkers that lead to potential future treatment of neurodegenerative diseases.

Identify Key Immunoproteins with LC-MS/MS

April 15, 2024

A recent study published in Molecules used our NanoLC-MS/MS platform to screen for plant-based proteins that could potentially boost the human immune response.

Colostrum, the milk produced immediately after birth, contains many proteins that transfer immunity to the newborn. The study screened a library of 100 plants to find proteins that behave like these transfer factors.

Researchers selected plant extracts based on a cytotoxicity assay evaluating their ability to enhance PBMC killing of human cancer cells, compared to that of the colostrum transfer factor. The two top candidates were Raphanus sativus and Brassica juncea. The 3-30 kDa ultrafiltrates of these two extracts, identified as having the highest PBMC cytotoxicity, were subject to NanoLC-MS/MS analysis.

Table 1: Top protein hits for Raphanus Sativus 3-30 kDa ultrafiltrate

The majority of identified proteins were seed storage proteins, with other proteins involved in plant defense and stress response. Notably, the analysis identified a defensin protein and a Kunitz trypsin inhibitor, both of which are involved in the plant immune response. This confirmed Raphanus sativus and Brassica juncea extracts as promising targets for further characterization of immune exploration, and potential treatments for humans using plant-based extracts.

Comments: The data here showed that our LCMSMS platform could reliably identify key target proteins present at low abundance. Our Mass Spectrometry facility has over 15 years of experience handling all sample types for applications ranging from protein identification & quantitation, PTMs to Host Cell Protein (HCP) related analysis. By employing top-of-the-line Orbitrap Exploris technology, we offer low cost, high sensitivity and high accuracy Mass Spectrometry analysis.

Anti-CHO HCP Antibodies with 99% Coverage

March 19, 2024

Looking for CHO HCP antibodies with high coverage?

Applied Biomics’ anti CHO HCP antibody offers ~99% coverage of many CHO strains, including CHO-S, CHO-K1, CHO-GS, and CHO-DG44. Our CHO antibodies detect over 3,000 low and high MW proteins, and come in both Serum and IgG form.

Our CHO antibodies have been repeatedly tested to ensure 99% coverage in each quadrant of the 2D Western Blot, including those low molecular weight HCPs. They are ideal for use in 1D/2D Western Blots, ELISA, and Immunoprecipitation (IP).

Want to test our antibodies first? As a leading provider of HCP antibody coverage services, we can test our antibody against your samples and compare with your in-house or other commercial HCP antibodies.

Get 20% off our antibodies when testing with our HCP Antibody Coverage service. Simply contact us and use the code ABO20HCP.

Left: 1D Western Blot showing CHO antibody coverage of various CHO strains
Right: Quadrant coverage of Anti-CHO HCP Antibody against CHO HCPs

Want to test our antibodies first? As a leading provider in HCP antibody coverage services, we can test our antibody against your sample and compare with your in-house or other commercial HCP antibodies.

Why us?

  • High HCP coverage: Each CHO antibody product has passed rigorous 2D Western Blot analysis to ensure overall HCP coverage > 99% and each quadrant coverage > 99%, including low molecular weight HCPs.
  • Competitive pricing: Same quantity at a lower price than similar antibodies from other vendors.
  • Quality guaranteed: We optimize our products to ensure consistency in performance matching the highest standards.
  • Excellent support: Backed by over 20 years of experience in antibody development and validation, our scientists will address your questions / concerns within 48 hours.

Achieve maximum HCP coverage with Applied Biomics antibodies

February 5, 2024

Frustrated with low antibody coverage in your HCP assays?

Applied Biomics’ new product website now offers HCP antibodies with well over 95% coverage of various host types!

We currently provide anti-CHOanti-E. coli, and anti-HEK293 antibodies in Serum or IgG form. Importantly, our anti-CHO HCP antibodies react well with CHO-S, CHO-K1, CHO-GS, CHO-DG44, etc. Our anti-HEK293 antibodies react well with not just HEK293, but other human cell lines such as CAP, Hela, Per.C6, MRC-5, etc. These antibodies are ideal for use in 1D/2D Western Blots, ELISA, and Immunoprecipitation (IP).

Left: Quadrant coverage of Anti-CHO HCP Antibody against CHO HCPs
Right: 2D Western Blot overlay of CHO HCP / Anti-CHO HCP Ab

Want to test our antibodies first? As a leading provider in HCP antibody coverage services, we can test our antibody against your sample and compare with your in-house or other commercial HCP antibodies.

Why us?

  • High HCP coverage: Each product has passed rigorous 2D Western Blot tests to ensure overall HCP coverage > 95% and each quadrant coverage > 90%, including low molecular weight HCPs.
  • Competitive pricing: Same quantity at a lower price than similar antibodies from other vendors.
  • Quality guaranteed: We optimize our products to ensure consistency in performance matching the highest standards.
  • Excellent support: Backed by over 20 years of experience in antibody development and validation, our scientists will address your questions / concerns within 48 hours.

New Year’s Promotion

January 3, 2024

New Year’s promotion: 10% Off Protein Identification (Offer Ends 02/03/2024)

Happy New Year to all! We would like to show our appreciation for choosing us to serve your proteomics needs. From January 3 to February 3, we are offering 10% off protein identification of 2D gel spots or 1D gel bands.

Take advantage of this limited-time discount to kick start your research for the new year. This offer applies to all our loyal existing customers as well as new customers.

*Terms: Order must contain at least 20 2D gel spots or at least 10 1D gel bands. Only applies for Protein ID by standard MALDI-TOF/TOF, not LC-MS/MS. Cost of running 2D DIGE gel not covered. Does not apply to previous orders. Applied Biomics’ standard service terms and conditions apply.

Evaluate Differential PTM Expression using 2D Western Blot

November 16, 2023

A recent paper published in PLOS One used our 2D Western Blot platform to study the function of S-Glutathionylation (SSG) levels in colorectal cancer samples.

The study design consisted of two 2D Western Blots and one 2D DIGE:

  • Gel-1 (2D WB): WT, Western Blot with Anti-Glutathione antibody
  • Gel-2 (2D WB): Tumor, Western Blot with Anti-Glutathione antibody
  • Gel-3 (2D DIGE): WT, Tumor

Gel-1 and 2 WB provided information about proteins with differential SSG levels in WT and Tumor, while Gel-3 provided information about protein ratios between WT and Tumor. After obtaining the protein ratio adjusted SSG ratios, several spots with the highest adjusted ratios were selected for Mass spectrometry analysis to obtain the protein identity and SSG sites in each protein.

Notably, the identified proteins include three glycolytic enzymes ( ENOA1, PGAM1 and PGK1) involved in diverting glycolysis & continuing glutathione synthesis. Taken as a whole, the 2D DIGE and Mass Spectrometry data provides insight into the pathogenesis of colorectal cancer and the role of SSG.

Comments: Post translational modifications (PTMs) play a key role in disease pathogenesis. However, the low level of PTMs makes it challenging to detect and quantify the PTM change in the complex proteome. This publications shows once again that our 2D Western Blot platform offers visualization, fast screening, and quantitation of PTM levels that no other platform can achieve. Further analysis by mass spectrometry can identify the protein PTM sites.

Identify Pancreatic Cancer Biomarkers with 2D DIGE

October 11, 2023

A recent study in Biomedicines used our 2D DIGE and Mass Spectrometry platform to identify potential biomarkers for pancreatic cancer.

The study focused on two kinase inhibitors, AT 9283 and WZ 3146, which had been previously identified as promising cancer targets due to their potency in inhibiting cell viability. A 2D DIGE experiment was run to compare WT pancreatic cancer cells with AT- and WZ-treated pancreatic cancer cells.

The 2D DIGE / Mass Spectrometry identified 60 proteins with differential expression between the WT cells and the two kinase-treated cells. The protein identities and ratios are summarized in the following heat map and table:

Notably, both lactoferrin and radixin were upregulated in the AT and WZ groups. Lactoferrin, is known to inhibit proliferation of cancer cells and induce apoptosis, while Radixin is involved in crosslinking PD-L1 to the actin cytoskeleton. PD-L1 is a significant antibody with a supposedly major role in suppressing autoimmune diseases.

Taken together, the proteomics and other data suggest treatment of pancreatic cancer involving PD-L1, and warrants further studies with AT and WZ.

Comments: The data here showed once again our 2D DIGE/MS platform could reliably identify key biomarkers that lead to potential future treatment of pancreatic cancer.

Effects of pyruvate kinase M (PKM2) Tyrosine Phosphorylation on bone cell differentiation

September 19, 2023

A publication in March 2023 used our nanoLC-MS/MS service to determine top candidates for phosphorylation. In the experiment, total cell lysate was prepared from IgSF11-treated culture, passed through phosphoprotein affinity columns, and separated by SDS-PAGE. After confirmation of tyrosine phosphorylation by Western Blot, the major phosphorylated band was excised for mass spectrometry analysis.

The nanoLC-MS/MS identified three candidates for phosphorylation: vimentin, tubulin beta 5, and PKM2. Importantly, data analysis identified phosphorylation at Y105 as a unique identifier of PKM2 in immunoglobulin superfamily 11 (IgSF11) activated proteins.

Further analysis showed that IgSF11 activates several tyrosine kinases, which phosphorylate PKM2, causing lower PKM2 activity. Conversely, IgSF11-deficient cells show higher PKM2 activity and defective osteoclast differentiation. The data indicated that PKM2 acts a metabolic switch in the development of bone cells.

Remarks: this publication shows that our mass spectrometry platform was able to identify a key phospho-protein as well as its specific phosphorylation site. More importantly, follow-up validation studies confirmed the significance of this protein and its phosphorylation in the context of bone cell differentiation.

Accurately detect and quantify Intact Protein Mass

August 17, 2023

Applied Biomics’ Intact Mass services offer accurate detection & quantitation of purified intact proteins. Using top-of-the-line Orbitrap Exploris technology, we have optimized from sample preparation to mass spectrometry scan settings for different kinds of proteins.

In an intact mass experiment, proteins are introduced into the mass spectrometer without digestion. Because each protein is different, mass spectrometry settings must be optimized. The mass spectrometer then separates the intact protein into components, producing a “charge envelope.” Typically there are multiple high-abundance m/z peaks, with the highest peak in the middle.

The raw data is then submitted to BioPharma Finder 5.1, which generates a “deconvoluted spectra.” The most abundant mass of the protein is shown as a dominant main peak, while other minor components are shown as smaller peaks. Both main and minor components will be quantified.

Data Report covers the following:

  1. MS Spectra: Protein fingerprint showing the peaks and charge states at various m/z
  2. Deconvoluted Spectra: Main peak & mass of the protein, along with smaller peaks of minor components
  3. Component-level data
  4. Charge state-level data

Accurately quantitate between samples with our LFQ service

July 10, 2023

Applied Biomics’ Label Free Quantitation (LFQ) services offer high-sensitivity detection and accurate quantitation of proteins in any protein sample. By combining optimized protocols and top-of-the-line Orbitrap Exploris Technology, we deliver high-quality data with fast turnaround and a low cost.

Label-free quantitation uses the intensities of spectra to compare relative & absolute abundance between samples. Due to the high sensitivity of Orbitrap Mass Spectrometers and sophisticated Proteome Discoverer software, this is a reliable approach with a more straightforward design than labeled methods.

The success of LFQ does not depend on labeling efficiency—a major challenge for labeled methods. In addition, LFQ is not constrained by the number of labeling reagents, allowing an unlimited number of samples to be compared. Finally, the protein coverage in LFQ is generally higher than in labeled approaches.

Our LFQ data report consists of the following:

  1. Protein abundance in each sample
  2. Confidence metrics
  3. Detailed protein information
  4. Protein functions, locations and pathways (if available in database)
  5. Peptide-level data

Phosphoprotein Profiling shows how CD44-dependent phosphorylation facilitates parasite invasion

June 14, 2023

CD44 is an important host factor for the invasion of malaria parasite but its mechanism is unknown. A recent study used our phosphoprotein profiling platform to compare both protein & Phosphoprotein profiles between WT RBCs and RBCs treated with Erythrocyte Binding Antigen-175 (EBA-175), a CD44 binding partner.

In the study, two sets of 2D DIGE experiments were performed:

  1. WT total protein vs EBA-175-treated total protein
  2. WT Phosphoprotein vs EBA-175-treated Phosphoprotein

The data revealed 50 spots with >1.3 fold protein level change, and 26 spots with >2 fold increased Phosphorylation in the EBA-175-treated sample. After the Phospho ratio was adjusted based on protein levels, the six spots with highest adjusted-Phospho ratios were identified by mass spectrometry.

Table showing the protein ID of six spots with highest adjusted-phospho ratios.

Taken as a whole, the proteomics data not only provides clues into potential proteins involved in the EBA-175 induced signaling pathway, but demonstrates that EBA-175 stimulation leads to increased Phosphorylation of several RBC proteins.

Comments:

Post translational modifications (PTMs) play a key role in cell signaling pathways. However, the low level of PTMs makes it challenging to detect and quantify the PTM change in the complex proteome. Our Phosphoprotein profiling platform offers: visualization, fast screening, and quantitation of phospho-protein changes that no other platform can achieve.

Analyze Time-Course Studies with 2D DIGE

May 22, 2023

A recent study in Blood Advances used our 2D DIGE service to analyze protein profile changes of platelets (PLT) over time.

Miyazawa B, Trivedi A, Vivona L, Lin M, Potter D, Nair A, Barry M, Cap AP, Pati S. Histone deacetylase-6 modulates the effects of 4°C platelets on vascular endothelial permeability. Blood Adv. 2023 Apr 11;7(7):1241-1257.

When blood is stored in blood banks, the characteristics of platelets may change based on time and storage conditions. In current practice, platelets are stored at 22 C for up to 7 days, but this may harm cell structure and function. Another storage method at 4 C reduces waste and better preserves function but decreases circulation time.

In the study, 2D DIGE followed by mass spectrometry was used to compare platelets from Day 1, Day 7 4 C, and Day 7 22C . The results showed 78 spots with differential protein & post-translational modification (PTM) levels between the storage conditions. Among them, 10 PLT proteins showed drastic difference at their PTM level. Notably, several tubulin isoforms were found to be significantly under-expressed at Day 7 at 4 C compared to 22 C. Consistently, authors showed that PLT α-tubulin acetylation is enhanced acutely by HDAC-6 inhibition.

Significant storage-induced changes in PLT posttranslational protein modification

The identified proteins represent potential biomarkers for storage-induced activation in platelets and provide clues into the mechanism by which HDAC-6 regulates platelet conditions.

Comments: The challenge of finding reliable biomarker is not only to identify changes at the protein level, but also at PTM level, and the hardest to confirm these findings using an independent approach. The data here showed once again our 2D DIGE/MS platform could fulfill all these objectives.

2D DIGE vs 2D Gel

April 17, 2023

2D gel electrophoresis is an effective method of separating protein components by PI (isoelectric point) in IEF (1st dimension) and molecular weight in SDS PAGE (2nd dimension).

2D DIGE is an improved version of standard 2D gel. Instead of staining after running the gel, 2D DIGE uses CyDyes to label up to 3 different samples prior to running the gel. This platform thus offers several advantages:

1. If using 2D gel to compare two samples, running two separate gels is required and there will be gel-to-gel variation. In 2D DIGE, because up to 3 samples are run on the same gel, this variation is eliminated and the cost is also lower.

2. CyDye labeling has a sensitivity 250x that of Coomassie and 5x that of silver staining. This allows extremely high accuracy, spot resolution, and reproducibility.

3. Standard size 2D gels sometimes lack proper separation and can overlook low-abundance proteins. Our 2D DIGE is large-format, allowing a wide dynamic range that can resolve over 5000 proteins and detect low-abundance proteins, large proteins and small peptides.

2D DIGE vs 2D Gel experimental procedure

Following are some applications of 2D DIGE that are impossible to achieve with 2D gel:

2023 New Year’s Promotion

January 3, 2023

Happy New Years 2023

New Year’s promotion: 10% Off Protein Identification (Offer Ends 02/03/2023)

Happy 2023! We would like to show our appreciation for choosing us to serve your proteomics needs. From January 3 to February 3, we are offering 10% off protein identification of 2D gel spots or 1D gel bands.

Take advantage of this limited-time discount to kick start your research for the new year. This offer applies to all our loyal existing customers as well as new customers.

*Terms: Only applies for Protein ID by standard MALDI-TOF/TOF, not LC-MS/MS. Cost of running 2D DIGE gel not covered. Does not apply to previous orders. Applied Biomics’ standard service terms and conditions apply.

News Release: Applied Biomics Offers Qualitative and Quantitative Host Cell Protein (HCP) Profiling Analysis

February 19, 2020

From Business Wire:

Applied Biomics, Inc., a leading Proteomics service provider, adds a HCP profiling analysis using their well established high resolution, large format 2-Dimensional Differential In-Gel Electrophoresis (2D DIGE) platform. This assay can be used to monitor HCP contamination during the purification process thus helps to improve purification strategy, specifically:

1) Visualize and overlay the HCP images between different samples
2) Quantitate HCP content by spot counting and PPM for each sample
3) Quantitate the purity of product or drug substance (DS) by spot counting and PPM
4) Quantitatively compare HCP content by fold-change and overall similarity between samples