ELISA ASSAY
Most of our experience analysing collected samples has been using allergen-specific ELISA immunoassays.
These cannot be used on the whole sample, rather the strip is retrospectively cut into small segments each corresponding to a period of individual observed activity. Each segment is then extracted and analysed separately. ELISA results are in units of pg of allergen per sampling time, where the time is determined for each segment.
We use the double-monoclonal allergen specific Der p 1 assay purchased from Indoor Biotechnologies (link). We modify this assay to increase its sensitivity about 500 fold; from approximately 2 ng/ml to 10 pg/ml and extract in volumes of 200 ul, thus giving a lower limit of detection of ~2 pg per sample.
The details of this assay have been published elsewhere (link). Also see the other PLoS papers for more minor details about performing ELISA's with samples. The 2013 paper is here, the 2016 paper is here.
It would also of course be possible to use other quantitative immunoassays. We have no experience of these. One example would be the state-of-the-art MARIA assays also from Indoor Biotechnologies. These are Multiplex assays performed on a Luminex xMAP® platform. The main advantages are they are quicker to perform, are highly sensitive and they can measure multiple allergens simultaneously on a single sample, however this comes at a cost.
Several sensor-based immunoassays for allergens have been reported more recently. We have no experience with these and don't know of any commercially available for indoor allergens. References, mainly from a recent review are here (link).
In the Development section we briefly discuss the possibility of developing a rapid immunoassay for the entire sample, (link)
There are pros and cons with ELISAs. The main strengths are the sensitivity of the assays and the quantitative nature of the results. The disadvantages are the extra handling and costs of analysing each segment. While (non-ELISA) visual-based assays in theory may provide additional information about the identity and size of each allergen-carrying particle, in reality it is hard to find a clinical interpretation for this information at present.
Most of our experience analysing collected samples has been using allergen-specific ELISA immunoassays.
These cannot be used on the whole sample, rather the strip is retrospectively cut into small segments each corresponding to a period of individual observed activity. Each segment is then extracted and analysed separately. ELISA results are in units of pg of allergen per sampling time, where the time is determined for each segment.
We use the double-monoclonal allergen specific Der p 1 assay purchased from Indoor Biotechnologies (link). We modify this assay to increase its sensitivity about 500 fold; from approximately 2 ng/ml to 10 pg/ml and extract in volumes of 200 ul, thus giving a lower limit of detection of ~2 pg per sample.
The details of this assay have been published elsewhere (link). Also see the other PLoS papers for more minor details about performing ELISA's with samples. The 2013 paper is here, the 2016 paper is here.
It would also of course be possible to use other quantitative immunoassays. We have no experience of these. One example would be the state-of-the-art MARIA assays also from Indoor Biotechnologies. These are Multiplex assays performed on a Luminex xMAP® platform. The main advantages are they are quicker to perform, are highly sensitive and they can measure multiple allergens simultaneously on a single sample, however this comes at a cost.
Several sensor-based immunoassays for allergens have been reported more recently. We have no experience with these and don't know of any commercially available for indoor allergens. References, mainly from a recent review are here (link).
In the Development section we briefly discuss the possibility of developing a rapid immunoassay for the entire sample, (link)
There are pros and cons with ELISAs. The main strengths are the sensitivity of the assays and the quantitative nature of the results. The disadvantages are the extra handling and costs of analysing each segment. While (non-ELISA) visual-based assays in theory may provide additional information about the identity and size of each allergen-carrying particle, in reality it is hard to find a clinical interpretation for this information at present.
CUTTING UP THE COLLECTION SURFACE
While we found workable ways to do this, I would not say the process was finalised and for each application and device some experimentation may be needed, depending on what materials are available. This however is a general guide as to what we did.
We have found it necessary to mount the collection surfaces (electret or iPad) onto another film to stabilise them for handing, cutting up and mounting onto the sampling wheel. We have used different double sided adhesives for this purpose, either the ones which come without a backing layer, (which don't seem to stick to themselves on the roll) and those which come with a removable backing layer (which is like silicone paper, nothing much will stick to it).
In some instances we have further bound this to a second plastic film (eg; over-head projector film) for ease of handling - we found this to particularly be the case with model 1, in order to keep the electret flat and not get stretched when removing from the wheel.
For cutting we have found it necessary to use a guillotine for strips or a punch for circles to be able to precisely cut 5 mm wide sections, (or 25 mm wide disks) - actually you want about 4.75 mm so they fit into the slot on the wheel.
In model 3, there is an indentation in the sampling wheel at the triangle position, so there is a region where the tapes can overlap. Use this as the start / stop region if possible.
After cutting on the guillotine, and before use, remove all excess backing layers and prior to sampling remove the top protective layer from the iPad adhesive. Inserting a tab while in the lab, prior to use, can be handy.
While we found workable ways to do this, I would not say the process was finalised and for each application and device some experimentation may be needed, depending on what materials are available. This however is a general guide as to what we did.
We have found it necessary to mount the collection surfaces (electret or iPad) onto another film to stabilise them for handing, cutting up and mounting onto the sampling wheel. We have used different double sided adhesives for this purpose, either the ones which come without a backing layer, (which don't seem to stick to themselves on the roll) and those which come with a removable backing layer (which is like silicone paper, nothing much will stick to it).
In some instances we have further bound this to a second plastic film (eg; over-head projector film) for ease of handling - we found this to particularly be the case with model 1, in order to keep the electret flat and not get stretched when removing from the wheel.
For cutting we have found it necessary to use a guillotine for strips or a punch for circles to be able to precisely cut 5 mm wide sections, (or 25 mm wide disks) - actually you want about 4.75 mm so they fit into the slot on the wheel.
In model 3, there is an indentation in the sampling wheel at the triangle position, so there is a region where the tapes can overlap. Use this as the start / stop region if possible.
After cutting on the guillotine, and before use, remove all excess backing layers and prior to sampling remove the top protective layer from the iPad adhesive. Inserting a tab while in the lab, prior to use, can be handy.