Label Free Quantitation
Our Label Free Quantitation (LFQ) services offer high sensitivity detection and accurate quantitation of proteins in any protein sample. By using the sum and intensity of spectra peaks to calculate abundance, this approach can quantify proteins between an unlimited number of samples without labeling. Applied Biomics’ LFQ services offer low cost, fast turnaround and high quality data by combining optimized protocols and top-of-the-line Orbitrap Exploris technology,
Main applications
- Study objectives: Comparing protein expression levels between experimental groups
- Sample type: All protein samples
We have optimized protein extraction for LFQ analysis on the following sample types:
- Whole cell lysate from cultured cells, bacteria and other organisms
- Subcellular organelles such as Mitochondria, Lysosome and Exosome etc
- Secretomes
- Protein complex such as IP or column purified complex
- Clinical samples with interfering substances such as high salt in urine
- Serum samples with high abundant proteins and a wide dynamic range
LFQ experimental procedure
- Sample preparation: Sample clean up, protein extraction, measure/adjust protein concentration, and trypsin digestion
- nanoLCMSMS: Peptides separated and analyzed by ThermoFisher Ultimate 3000 & Orbitrap Exploris
- Data analysis: Protein identification and Label Free Quantitative Proteomics analysis by Proteome Discoverer
- Data Report covers the following:
- Abundance data for proteins in each sample and protein ratios
- Confidence metrics
- Detailed protein info
- Protein functions (if available in database)
- Peptide-level data
Label Free Quantitation vs. Label-Based Quantitation:
Label Free Quantitative Proteomics uses the signal intensity of ions to quantify the protein abundance. In contrast, Label-based methods, such as iTRAQ or TMT, labels the peptides and protein with stable isotopes. thus allowing quantitative comparisons between labeled samples. Please view the following table to guide your decision:
| LFQ | iTRAQ/TMT | |
|---|---|---|
| Sample type | All protein samples | All protein samples |
| Number of samples | Unlimited | Limited based on the kit |
| Sample preparation | Samples are processed individually | Samples are processed, labeled individually, then combined |
| NanoLC-MS/MS | 1 run per sample | 1 run per project |
| Protein coverage | Higher | Lower |
| Main drawbacks Solution | Run-to-run variation Run triplicates | Labelling efficiency Fractionate protein |
In-solution Samples
Sample type: We accept in-solution, in-gel samples or IP beads for this service.
Protein amount: 5-200 µg of total protein from each sample. For 1D gel bands, 5-20 ng protein / band is preferred.
Protein concentration: 5-20 mg/ml is preferred for in-solution samples.
Buffer: Please feel free to submit samples in whichever buffer that you would normally use. Please inform our scientists about buffer composition. Additional charge may apply on samples that require extensive amount of the extra work such as protein eluting, concentrating, buffer exchange or serum abundant protein depletion.
Biohazardous Materials: We do NOT accept samples containing any biohazardous materials. If your samples contain any biohazardous materials, please make sure to deactivate them completely using the appropriate procedure and declare them in the order form. In addition, we do NOT accept any samples containing Level 4 biohazardous materials even if they are deactivated. Please view here for the rank of biohazardous materials.
Sample type: Please inform our scientists about your sample type. We will send you additional tips for each sample type.
Pricing
| Services Description | Academic/Governmental Labs | Industrial/Commercial Labs | ||
|---|---|---|---|---|
| Code | Price | Code | Price | |
| Protein Fractionation | 355A | call for price | 355N | call for price |
| LC-MS/MS on Low Complexity Samples (1D Gel band, 2-5 samples) | 331A | $499/sample | 331N | $589/sample |
| LC-MS/MS on Low Complexity Samples (1D Gel band, 6 or more samples) | 331A | $429/sample | 331N | $499/sample |
| LC-MS/MS on Moderate Complexity Samples | 332A | call for price | 332N | call for price |
| LC-MS/MS on Moderate to High Complexity Samples | 334A | call for price | 334N | call for price |
| LC-MS/MS on High Complexity Samples | 333A | call for price | 333N | call for price |
Each order should have a minimum of 2 samples. Turnaround time is ~7 business days.
Sample category:
- Low complexity samples: 1D gel band – Each sample should contain only one gel piece with one protein band, no bigger than 1x1x5mm in size, <10 proteins. Samples with more than one gel piece, and/or more than one protein band will be considered as moderate to high complex samples. We recommend you cut out only the area with your protein of interest (1x1x2mm size) and follow these guidelines to prepare samples
- Moderate complexity samples: protein mixture such as column purified sample, <100 proteins
- Moderate to High complexity samples: Protein mixtures such as subcellular organelles, IP samples
- High complexity samples: whole cell lysate, tissue, and others
Price covers the following:
- NanoLC-MSMS
- Standard database search against one species in NCBI or SwissProt database using SequestHT
Price does NOT cover:
- Sample preparation
- Data request in addition to the standard report