Archive for December 2021

Inside the Lucira Check It COVID-19 Test

December 3, 2021, 5:00 pm

This post was previously published on the substack.

The Lucira COVID-19 test is an “at-home” LAMP qPCR-like test. Essentially you receive disposable molecular test device, with embedded reagents for $55 which you use once and then throw away.

I was curious to better understand the instrument and went hunting for teardown pictures and patents. Luckily Brad Ackerman on pulled one apart and posted pictures on twitter. Some of these are included here with permission from Brad. The teardown pictures also closely match one of Lucira’s patents which gives us another source of information. The patent has some wonderful figures, and gives us a really nice exploded view of the device:

This really gives us a great overview of the instrument. We can see a single main PCB with contains LEDs, a photosensor, heating element, temperature sensor and microcontroller. The heating element is a simple PCB trace used for resistive heating. This is required get the reaction to ~60C as required for LAMP. Critically, for LAMP we don’t need do thermo-cycle, so cooling is not required. 

The photosensor sits in the center of the heating element, surrounded by LEDs. On top of this there’s a reasonably complex optical system:

The sample will flow down from the prep tube and be exposed to a reagents ending up in a number of fluidic chambers. This is kind of neat, because as we can see in the image above each LED is paired with a chamber. The light from the LED is reflected through the chamber and down on to a single photosensor.

This allows you to multiplex to some degree, using a single sensor and multiple LEDs which you can switch on/off. Naively you might think that for COVID-19 you only need a single measurement. But the FDA authorization makes it clear that the device also has a “Positive Internal Control (PIC) and Lysis Internal Control (LIC)” which must also show the expected results.

Looking at Brad’s teardown images, we can see that Lucira have only placed 4 of the possible 8 LEDs in the COVID-19 instrument. So Lucira could potentially expand the capability of the device to other targets/variants:

I wasn’t able to identify the photosensor used here, but I suspect it doesn’t cost more than $1, similarly the LEDs likely cost almost nothing. The microcontroller is an STM32F030C6, a simple 32bit ARM microcontroller with 32K of flash, 4K of RAM. This also usually costs ~$1. Overall I suspect the BOM cost is somewhere in the region of $5. The photosensor could potentially be the most expensive part if this turns out to be a high sensitivity photodiode…

The optical system appears to be all plastic, here you can see the light pipes. They show up as different colors, it’s possible there are embedded filters. You can also see the heat conductive paste that is used to couple the heating element to the rest of the unit:

The result of all this is a platform, where much like qPCR, you can monitor the amplification process in order to determine the presence/absence of your target. The patent shows some example experimental data:

While I suspect the electronics doesn’t cost much, I’m still impressed they can sell this for $55, as it’s a reasonably complex optical/reagent system. I’d be curious to know what their margins are. Overall I find the approach pretty neat. No doubt BARDA (who gave them $21.9M in 2018) must consider this a prescient funding decision.

Lucira IPO’d in February, but it seems that their share price has been rather unstable. It will be interesting to see what happens with Lucira and if they and gain adoption outside of COVID19 diagnostics.

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Roswell Revisited

December 2, 2021, 5:00 pm

This post previously appeared on the substack.

I’ve previously talked about Roswell. But they’ve recently been making a bit more noise, and I thought It would be interesting to review my previous post in light of this new information.

Roswell was founded in 2014, back in 2018 they were reported to have raised $6M. Cruchbase now lists them as having raised $37M though it’s not clear how recent this figure is. A recent Genomeweb article says they now have 50 employees.

Roswell’s patents show a basic configuration where a strand of dsDNA is suspended between two electrodes as a kind of scaffold. A polymerase is then attached to this scaffold. The polymerase performs synthesis of a template strand and conformational (and other changes) are detected:

Detection is likely through tunneling (and potentially other) currents between the electrodes. Conformational changes in the polymerase and/or incorporated nucleotides would result in current changes that could be detect to sequence the strand under synthesis.

The patent I previously looked at showed data from homopolymers, with current defections on the order of 60pA.

So what’s new? Well, if the animations on their website are anything to go by, the basic mechanism is still broadly as described above:

Roswell’s whitepapers also help clarify this process, where an electric field is used to attract the bridge molecule (dsDNA). The approach seems pretty neat, as you can turn on the field, attract and attach the dsDNA and then immediately turn off the field. This means that you should be able to attach a single bridge molecule to each sensor. Without something like this, you would be Poisson limited, with many sensors having no, or multiple bridges.

The Genomeweb article suggests they’re shifting away from DNA sequencing and “targeting diagnostics, especially for infectious diseases, or drug discovery as its first applications”. This is perhaps because their sequencing accuracy isn’t quite competitive yet:

“”Discrimination of the four bases can approach over 90 percent, and in complex templates, 70 percent,” Mola said.”

It’s difficult to unpack this statement without knowing exactly what experiments were performed and how the data was processed. But to me it suggests they’re not quite sequencing yet, but rather looking at ensemble properties of synthetic templates.

The white paper also tends to focus on applications other than DNA sequencing. For example hybridization based assays:

I personally find these other applications less compelling than sequencing. The advantage over other approaches, including other single molecule approaches, is unclear to me. But I do find the fact that the approach works at all, kind of exciting.

The other nice feature of the Roswell approach is that they can potentially pack features much closer than you can on a protein nanopore chip. The density they show, of 2500 sensors per mm² seems pretty good.

Roswell Gen3 chip: 180nm process, 4x6mm die, 1KSPS (per sensor)

This is an advantage because it keeps the chips relatively small. Roswell’s Gen3 chips are 4x6mm. When I last looked at image sensor pricing, it seems like chips of this size would generally retail for about $10.

That’s pretty cheap, but still an expensive COGS for a consumable, which given the approach I suspect is not washable/reusable. This potentially makes the approach unsuitable for low-cost diagnostic applications.

However, that their approach works at all is neat, and I’ll be watching Roswell’s progress with interest.

Disclaimer: As always, you should be aware that I have equity in sequencing companies (based on prior employment) and am current working on a novel sequencing approach.

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Bio-rad SmartSpec Plus Spectrophotometer

December 2, 2021, 2:30 am

I picked up a Bio-rad SmartSpec UV-Vis Spectrophotometer on eBay. The SmartSpec uses a Xenon flash lamp and is a much more compact/similar design than the DU530/DR4000 I previously looked at. Of course I decided to pull it apart, and my pictures are below for reference.

The basic layout is shown below. The PCB looks like a very traditional 1980s/90s design:

The lamp sits above the spectrometer block in the metal enclosure, with the cuvette sitting between lamp and the spectrometer:

Inside the spectrometer block we see a couple of mirrors and the diffraction mirror:

It uses a traditional design, fairly similar to that employed by Ocean optics and other spectrometer makers:

The Ocean optics block, from here.

The light essentially reflects off the diffraction mirror and into a linear CCD. I suspect this is an ILX511, which is pretty common in spectrometers and I’ve played with before.

A few more pictures of other parts of the instrument are below. I’ll add more posts if I do any more work with this device.

Mirror
Diffraction Mirror
CCD
Power Supply
Back of the CCD PCB
The printer, mine is broken.
Another printer image, showing the interface board and FPCB.

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AxBio – Nanopore Sequencing Startup

December 1, 2021, 5:00 pm

This post previously appeared on the substack.

AxBio was founded in 2016 by Hui Tian and Igor Ivanov. Hui was previously Vice President of Genia Technologies. The company appears to be funded by Tsingyuan (now Foothill) Ventures, who interestingly also invested in Quantapore. 

AxBio appear to be developing a “forth generation” DNA sequencing platform, which they say can “can shorten the human genome sequencing from the current 1-2 weeks to one day”. 

In addition to this, they (surprisingly to me) also sell a number of generic extraction kits:

It’s hard to find out much about AxBio, but they suggest they have a 1 million sensor chip. Other reports suggest this is a “nanopore sequencing by DNA synthesis”..approach using a…“hydrodynamic chip combining a silicon CMOS base and a biological membrane”… “together with a polymerase and labeled nucleotides”.

This kind of suggests an approach similar to Genia, where tagged nucleotides are incorporated and read in-situ:

Genia’s Nanopore-SBS approach

Unfortunately, I’ve not seen much from Genia/Roche since I last wrote about them in 2016. Suggesting progressing the platform to a point where it’s competitive is proving problematic.

AxBio’s patents suggest a similar approach, where a polymerase is used in-situ to synthesis a strand containing labeled nucleotides.

Synthesizing in-situ may help slow the translocation of the strand through the pore, which would generally be required for any ionic current detection method. They also seem to suggest reading the same strand multiple times, using rolling circle amplification to create multiple copies of the input DNA.

This nanopore-SBS approach has the obvious disadvantage that you are not sequencing the original material and as such will not see base modifications, or be able to natively sequence RNA.

As well as protein nanopores, Axbio patents discuss solid state nanopores at length which suggests a different sensing mechanism is also being considered. In particular the patent states that impedance sensing may be used to sequence by “detect one or more signals indicative of an impedance or impedance change in the nanopore when the at least portion of the tag is within the nanopore”.

This sounds pretty ambitious. A single tag would need to create an impedance change within the volume of the nanopore which can be detected during the short period of time it resides within the pore. In addition to this, you will likely have impedance contributions from multiple tags, background impedance contributions from the buffer, and potentially if the electrodes a close enough, tunneling currents between the electrodes.

The patents don’t contain any example procedures or datasets that I could see. So I went looking for support for impedance sensing approaches in the literature. My friend Shohei manage to find a publication where a single molecule induces a capacitance change by displacing a membrane. But this is a big leap from detecting impedance changes created by single molecules, and we were unable to find much support in the literature for this sensing method.

Outside of nanopores, scanning capacitance microscopy is a relatively established probe microscopy technique. In this article exploring the limits of SCM, they suggest that best resolution you can obtain in the Z direction is 1nm, and 5nm in XY. Here using a bandwidth of ~1KHz. For the most part SCM appears to be used for imaging semiconductors and similar planar surfaces. This is clearly a lot easier than measuring single molecule tags.

5nm would suggest we would see contributions from ~15 bases/tags. Which would be significantly worse than the ~5 you see with protein nanopores.

So overall, the impedance based approach seems like it would be really hard to get working.

AxBio however appear to suggest that they have “released the latest single-molecule gene sequencer” so it will be interesting to see what actually appears. They talk about a 1 million sensor thumb nail sized chip. This also suggests a solid state approach, as achieving this density with protein nanopores is likely challenging. So, I remain curious as to what will actually appear and will be keeping an eye out for any data releases from AxBio.

Disclaimer: As always, you should be aware that I have equity in sequencing companies (based on prior employment) and am current working on a novel sequencing approach.

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