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Short answer
The micro BCA assay is the bicinchoninic acid protein assay in a format built for dilute samples. In Thermo Fisher’s Micro BCA Protein Assay (Cat. 23235) the working range is 2-40 µg/mL with a 150 µL sample, or 0.5-20 µg/mL with a 500 µL sample, read at 562 nm after 60 minutes at 60 °C. The standard BCA kit covers 20-2,000 µg/mL.
What the micro BCA assay is
It uses the same chemistry as the standard BCA protein assay, with a reagent kit adapted for low concentrations. Thermo Fisher describes its micro kit as a “special 3-component version” of its BCA reagent kit for dilute protein solutions. The standard kit it compares against is a two-component system.
The chemistry comes from Smith and colleagues (Analytical Biochemistry, 1985). Protein reduces Cu2+ to Cu1+ in alkaline solution. Bicinchoninic acid then forms a purple complex with the cuprous ion, and the colour rises in proportion to protein over a broad range. The authors reported that the method tolerates nonionic detergents and simple buffer salts better than the Lowry method, while keeping the Lowry method’s high sensitivity and low protein-to-protein variation.
How the micro format differs in practice
The micro format changes the working range, the sample volume and the incubation, not the readout. Both formats in Thermo Fisher’s comparison are read at 562 nm, so the same plate reader or UV-Vis spectrophotometer works for either; a plate reader is the usual instrument for microplate work.
Micro BCA versus standard BCA
Every value below comes from Thermo Fisher’s BCA protein assays page, which compares its Micro BCA Protein Assay (23235) with its standard BCA kit with dilution-free BSA standards.
| Parameter | Micro BCA (23235) | Standard BCA |
|---|---|---|
| Working range | 2-40 µg/mL (150 µL sample) or 0.5-20 µg/mL (500 µL sample) | 20-2,000 µg/mL (25 µL sample) |
| Incubation | 60 min at 60 °C | 30 min at 37 °C |
| Total assay time | 130 min | 45 min |
| Absorbance | 562 nm | 562 nm |
| Reagent kit | 3-component | 2-component |
| Dilution-free BSA standards included | No | Yes |
| Detergents | Compatible | Compatible |
| Reducing agents and chelators | Incompatible | Incompatible |
The table shows the cost of the extra sensitivity. The micro format is about three times longer end to end (130 versus 45 minutes), uses a hotter incubation, takes a larger sample volume and does not supply ready-made standards. The two ranges barely overlap: the micro range tops out at 40 µg/mL, while the standard range starts at 20 µg/mL.
Where it matters for labs
Choosing between the two formats
The table suggests a simple screen based on the expected concentration of the sample.
- If the sample is expected to fall between 20 and 2,000 µg/mL, the standard kit’s range covers it with a 25 µL sample and a 45-minute assay.
- If the sample is dilute enough to sit under about 40 µg/mL, the micro kit’s two volume tiers apply: 2-40 µg/mL with 150 µL, or 0.5-20 µg/mL with 500 µL. The 500 µL tier reaches lower concentrations but uses more sample.
- Because the micro kit ships without dilution-free standards, a standard series has to be prepared in the lab, which belongs in the time budget. The standard-curve method is covered in the BCA standard curve guide.
Screening the sample buffer first
Both formats are listed as compatible with detergents and incompatible with reducing agents and chelators. With a dilute and often scarce sample, check the buffer composition against that list before committing it to the assay. If the buffer contains a reducing agent or a chelator, the assay is not a suitable fit as listed, and the Bradford assay works on a different principle and may be worth comparing. Thermo Fisher also lists less protein-to-protein variation for BCA than for Bradford. Upstream steps such as protein purification are where dilute samples and buffer additives usually originate.
Common misunderstandings
“Micro BCA is just the standard assay with less sample.”
The micro format uses a different reagent kit (3-component against 2-component), a different incubation (60 °C against 37 °C) and a larger sample volume, not a smaller one.
“A higher temperature means a faster assay.”
Not here. The micro kit incubates hotter but its total assay time in Thermo Fisher’s comparison is 130 minutes, against 45 minutes for the standard kit.
“Any buffer that works in the standard assay is fine at low concentration.”
The compatibility listing is the same for both formats, so a detergent-tolerant buffer is fine, and a reducing agent or chelator is a problem in either. Dilute samples make interferents more important to rule out, because there is less protein signal to spare.
FAQ
What is the micro BCA protein assay range?
For Thermo Fisher’s Micro BCA Protein Assay, 2-40 µg/mL with a 150 µL sample or 0.5-20 µg/mL with a 500 µL sample.
What is the micro BCA assay protocol?
The published parameters for Thermo Fisher’s kit are a 150 µL or 500 µL sample, a 3-component reagent, 60 minutes at 60 °C, a reading at 562 nm and a 130-minute total assay time, with the standard series prepared by the user. The step-by-step procedure belongs to the manufacturer’s instructions for use for the kit in hand, which is the document to follow at the bench.
When should I use micro BCA instead of standard BCA?
When the sample is expected to fall below the standard kit’s 20 µg/mL lower limit, within the micro kit’s 0.5-40 µg/mL span.
Can I use micro BCA with detergents or reducing agents?
Detergents are listed as compatible. Reducing agents and chelators are listed as incompatible.
Sources
- Thermo Fisher Scientific, BCA protein assays (URL checked 2026-10-29): Micro BCA Protein Assay, Cat. 23235, compared with the standard BCA kit.
- Smith PK, et al. Measurement of protein using bicinchoninic acid. Analytical Biochemistry 1985;150(1):76-85. PMID 3843705 (abstract checked 2026-10-29).
