Cellular Assays

Sample submission templates (email to Honghong Sun or Yangzhu Du):

• Luminex sample list
• Digital ELISA sample list_34
• Digital ELISA sample list_38
• Digital ELISA sample list_86

Existing Luminex  or other cellular assay projects billed through PBR, click here to submit samples online (PennKey login required).  Existing projects billed through iLab, click here to submit samples online (login required). 

The HIC offers turnkey service for a range of cellular immunology assays:

• Standard or digital ELISA (also called simoa, single-molecule assay) using Quanterix - Simoa® assay kits
• Luminex multiplex assays using off-the-shelf or custom panels
• T cell, B cell ELISPOT; plate reading/photo service (sorry, we do not offer read-your-own - see Cellular Assays FAQ below for reasons)
• Multicolor immunophenotyping using off-the-shelf or custom stain panels
• Consultation on assay design, selection, quotations
• Letters of support, methods, budgeting, or other information for grant proposals

Prices for cellular assays are shown on the full price list, downloadable under the Accounts, Billing, and Fees section, and a downloadable pdf of prices is also available (PennKey login required). Request a consultation if you would like a price quote.

STEP 1: Request a consultation and/or quotation by contacting Acting Technical Director Dr. Honghong Sun, PhD. Neither a PennKey nor a HIC account is required to schedule a consulation. Please be prepared with

• the overall goal/question and projected timeline of your experiment
• the assay or service you would like (Simoa, Luminex, etc.) - Dr Sun can assist with assay choice
• sample type(s) including species if non-human, and approximate number
• desired stain panel, analytes, use of off-the-shelf or custom stains / kits

Bring any questions or thoughts you may have on experimental design, customizing workflows for downstream applications, SOPs, etc. The HIC runs assays on a first come - first serve policy, so please allow 6-8 weeks of lead time as you plan your project with us. 

STEP 2:  Once you determine to do a project with the HIC, Path BioResource (PBR) staff will work with you to set up your study or project. Contact PBRHELPDESK@pennmedicine.upenn.edu, CC Dr. Honghong Sun. Download a copy of HIC Accounts & Billing Workflows. or visit our Accounts, Billing, and Fees section for instructions on how to set up or update HIC accounts.

STEP 3:  Submit samples online and ship or drop off physical samples to SCL 412.

STEP 1:  Download the appropriate sample list template from the following list, fill it in, and email to Honghong Sun at hsun2@pennmedicine.upenn.edu or Yangzhu Du at Yangzhu.Du@pennmedicine.upenn.edu. A list of samples must be received before your assay can be scheduled.  The HIC runs assays on a first come - first serve policy, so please allow 4-6 weeks of lead time.

• Luminex sample list
• Digital ELISA sample list_34
• Digital ELISA sample list_38
• Digital ELISA sample list_86

Sample types such as cell culture supernatant, body fluid, cells, or other:  Please list your samples in an Excel spreadsheet, provide any information requested during your project consultation (such as sample volume or dilution), and email to Honghong Sun.

STEP 2:  Submit an online request.  For Luminex or other cellular assay projects, click here to submit samples online (PennKey login required). Select your project and note that your sample list has been emailed to Yangzhu Du or Honghong Sun - it is not necessary to re-enter each individual sample.   Click here if your project will be billed through iLab (login required). 

Review our sample preparation guidelines and submission requirements if not already done.

STEP 3:  Drop off samples at Stellar-Chance Labs (SCL) room 412, or ship them to this address.  Samples received outside of our posted cut-off times require approval at least 24 hours in advance of receipt and additional charges will apply.

Contact Honghong Sun Cc Yangzhu Du with any questions.  The HIC shipping address is Attn: Yangzhu Du, University of Pennsylvania, Stellar Chance Labs Rm. 412, Philadelphia, PA 19104.  Telephone: +1 215-573-5367.  Email: Yangzhu.Du@pennmedicine.upenn.edu

Download our sample preparation guidelines and submission requirements. We accept serum, plasma, cell culture supernatant, and body fluids for Luminex, standard ELISA, or digital ELISA (Simoa) assays.  We also accept cell samples for flow cytometry or ELISPOT assays.

Sample preparation considerations are critical for the success of your assay. For example, for cytokine assays immediate processing of the blood sample is very important because many cytokines have a very short half-life. Refrigerate the tube immediately after a blood draw if you are in a pinch and can't get the sample to the HIC right away. Please consult with Dr. Honghong Sun in advance.

Plasma sample preparation

• Use EDTA tubes rather than heparin tubes, since heparin may interfere with Luminex readings
• Use harder spins if you don't care about the cells (up to 1300g for 10 minutes) or ask Dr. Honghong Sun for guidance

Serum sample preparation

• Hold whole blood at room temperature before spinning; if you fail to do this, the sample may re-clot and interfere with your assay
• Hold serum separator tubes (SST, also known as gold-top or marble-top) at room temperature for 30 minutes, then centrifuge (1000g 15 minutes) to remove clotted material before withdrawing serum
• Hold red-top tubes at room temperature for 60 minutes, then centrifuge (1000g 15 minutes) to remove clotted material before withdrawing serum

Culture supernatant, body fluids – as determined in your project consultation with Dr. Sun. 

Cell samples for flow cytometry assays - as determined in your project consultation with Dr. Sun. 

Sample submission requirements

• Clearly label sample container(s) with your name, the date, and HIC project name or number.
• Samples must be in Eppendorf tubes. Cell culture or assay plates are not accepted.
• For samples other than plasma or serum, please provide 10 mL/plate of the culture medium or solution in which your samples are dissolved.
• DO NOT THAW frozen samples! Please deliver frozen samples to the HIC (412 SCL) on dry ice.
• Don't forget to include internal control samples if available / appropriate for your experiment.
• If you know your sample dilution factor, please indicate this on the sample list template. If not, the HIC will follow protocol instructions for dilutions (unless arranged otherwise in advance).
• Please submit sufficient sample volume for duplicating your test (refer to kit for recommended volumes):
  • 65-120 uL for Luminex assays
  • 100-250 uL for digital ELISAs

Any leftover samples will be returned to you or discarded with your permission.

Download sample list forms above under Submit and Deliver Samples.

Questions about sample prep, completing sample list, sample delivery?  Please email Dr. Honghong Sun at hsun2@pennmedicine.upenn.edu or call her office at 215-573-7389.

HIC Shipping Address  Attn: Yangzhu Du, Stellar Chance Labs Rm. 412, 422 Curie Blvd., Philadelphia, PA 19104. Phone # 267-764-6703 and email Yangzhu.Du@pennmedicine.upenn.edu

What are Simoa (digital ELISA), Luminex, and standard ELISA assays?

Simoa stands for SIngle MOlecule Assay.  It is also called digital ELISA.  It is a single-molecule enzyme-linked immunosorbent assay that can detect proteins at sub-femtomolar concentrations. Thus, it is an ultra-sensitive immunoassay technology that allows detection of proteins and nucleic acids at very low levels (fg/mL).  This simple, reliable, and sensitive immunoassay platform measures biomarkers in most common sample types.

Luminex is an immunoassay that precisely measures multiple analytes in one sample.  It is a bead-based immunoassay that allows for multiplex detection of up to 100 analytes simultaneously.  Color-coded beads, or microspheres, are internally dyed with different proportions of red and infrared fluorophores that correspond to a distinct spectral signature or bead region. The quantification of multiple cytokines and other biomarkers in a sample provides critical information about biological processes and diseases.

ELISA is short for Enzyme-Linked ImmunoSorbent Assay.  ELISA is a technique to detect the presence of a single antigen in biological samples. ELISAs, like other immunoassays, rely on antibodies to detect a target antigen via highly specific antibody-antigen interactions. In a standard ELISA assay, the plate is first coated with an antibody specific to the antigen of interest.  The antigen in biological samples is captured through the antibody-antigen interaction. The antigen is then complexed to a detection antibody that is conjugated to biotin or streptavidin-HRP.  The signal generated can be detected by colorimetric or chemiluminescent methods. 

When is a simoa assay chosen?

Some analytes are present in samples at a level below the limit of detection of standard ELISA or Luminex.  Simoa allows detection of proteins and nucleic acids present at very low levels (fg/ml) and/or low-volume samples.

The typical detection range for a digital ELISA is 0.1 to 1 fmole or 0.01 to 0.1 ng, with sensitivity dependent upon the particular characteristics of the antibody-antigen interaction.  For most Luminex analytes, we observe a limit of detection of 1-10 pg/mL using cytokine standards diluted in a serum standard buffer.

Why doesn't the HIC allow investigators to scan their own ELISPOT plates? 

We have learned that the best way for many different labs to benefit from the scanner is to have trained HIC staff operating it.  We are experienced with "calibrating" the instrument when it comes to overlapping spots, very small or fuzzy spots versus artifacts, unequal spot distributions within a well, etc.  The HIC does not have the manpower to train others, help them troubleshoot, or manage instrument scheduling.  Moreover, many clients have found it faster and less expensive in terms of their trainee/staff time to have the HIC scan multiple ELISPOT plates for larger experiments.