cfDNA vs. ctDNA: Finding the Intruder in the House

One of the most confusing aspects of liquid biopsy is the terminology. You will hear "cfDNA" and "ctDNA" used almost interchangeably, but in oncology, the distinction is everything.

Think of the bloodstream as a crowded house party.

cfDNA (Cell-Free DNA): This represents everyone* at the party. It is the total count of DNA fragments floating in the blood. Most of these guests are normal—DNA from dying white blood cells, gut lining, and endothelial cells.
ctDNA (Circulating Tumor DNA): These are the intruders*. This is the specific fraction of DNA that came from a cancer cell.

The Challenge: Tumor Fraction

In a dog with cancer, the ctDNA is often a tiny minority of the total cfDNA.

* The "Tumor Fraction": In early-stage cancer, ctDNA might make up only 0.1% to 1% of the total cfDNA.
The Problem: Standard methods (like a Qubit fluorometer) measure the total* cfDNA.

Example 1: The False Negative
A dog has a small bladder tumor (TCC). The tumor is shedding DNA, but the dog is otherwise healthy.
* Total cfDNA: 0.8 ng/mL (Normal Range).
* ctDNA: 0.008 ng/mL (Present, but hidden).
Result:* A test measuring only "concentration" will say this dog is normal. You need a test that sequences the DNA to find the specific BRAF mutation hidden in that 0.8 ng/mL.

Example 2: The False Positive
A dog has severe dental disease and systemic inflammation.
* Total cfDNA: 5.0 ng/mL (Very High).
* ctDNA: 0.0 ng/mL (No cancer).
Result:* A test measuring only "concentration" will flag this dog as "High Risk," causing panic. But a sequencing test will see that all the DNA is normal, wild-type canine DNA. There is no cancer, just inflammation.

When is Total cfDNA useful?

Total cfDNA (concentration) is useful as a non-specific marker of "illness burden" (trauma, sepsis, necrosis).

However, for cancer detection, we need Tumor-Specific assays. These look for:
1. Mutations: Changes in the code (e.g., BRAF, c-KIT, TP53).
2. Copy Number Variations (CNVs): Chunks of chromosomes that are duplicated or deleted.
3. Methylation: Abnormal chemical tags on the DNA.

Conclusion

* cfDNA is the haystack.
* ctDNA is the needle.

Don't just weigh the haystack; use technology that can find the needle.