Company: BLLN
Filing Date: 2025-10-07
Form Type: S-1
Source: 0001193125-25-233697
Chunk: 195

Company: BillionToOne, Inc.
Filing Date: 2025-10-07
Form: S-1
Chunk 195
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 screening. Hum Genomics. (2019);13(1):62. |

| 45 |     | Dang, D. K., & Park, B. H. (2022). Circulating tumor DNA: current challenges for clinical utility. Journal of Clinical Investigation, 132(12), e154941 |

| 46 |     | Bronkhorst, A. J., & Holdenrieder, S. (2023). The changing face of circulating tumor DNA (ctDNA) profiling: Factors that shape the landscape of methodologies, technologies, and commercialization. Medizinische 
 Genetik, 35(4), 201-235.                                                                                                                                                                                         |

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even for relative quantification. Moreover, these NGS methods struggle with the easiest copy number analyses, such as the ability to distinguish one copy of a gene versus two copies, as may be
needed for a standard germline or carrier testing. Consequently, the gold standard for clinical testing of copy number analysis is pre-NGS technologies, such as microarray and multiplex ligation-dependent
probe amplification. The identification of these copy number changes in cfDNA requires detecting a change that is more than 100 times smaller, since the fraction of cfDNA that is derived from the fetus or the tumor can constitute less than 1% of the
total cfDNA. Only our smNGS platform can detect CNVs in cfDNA at these levels today. Similarly, smNGS is needed for determining fetal risk in single-gene recessive conditions and precisely quantifying response to therapy, as these problems require
absolute quantification of a low cfDNA signal against a high background originating from other tissues.

Precision of measurement 1% BillionToOne's smNGS encoding ddPCR 10% 100% 1 10 100 NGS 1000+ Number of loci assayable

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The components of our platform

Our smNGS platform seamlessly integrates several tools and patented technologies, as depicted in the figure below.

Quantitative Counting TemplatesTM (QCTSTM)* Specialized & scalable infrastructure Our smNGS platform is protected by an expanding moat. Custom bioinformatics with machine learning Engineering biology approach 50+ issued patents** & knowhow * patented ** as of Jun 30, 2025

Quantitative Counting Templates (QCTs)

QCTs are the foundation of our smNGS
technology. They are artificial DNA fragments that we design and synthesize to mimic the properties